Biosensors for biological or chemical analysis and methods of manufacturing the same

ABSTRACT

Biosensor including a device base having a sensor array of light sensors and a guide array of light guides. The light guides have input regions that are configured to receive excitation light and light emissions generated by biological or chemical substances. The light guides extend into the device base toward corresponding light sensors and have a filter material. The device base includes device circuitry electrically coupled to the light sensors and configured to transmit data signals. A passivation layer extends over the device base and forms an array of reaction recesses above the light guides. The biosensor also includes peripheral crosstalk shields that at least partially surround corresponding light guides of the guide array to reduce optical crosstalk between adjacent light sensors.

CROSS-REFERENCE TO RELATED APPLICATIONS

The present application is a divisional of U.S. application Ser. No.15/175,489, filed on Jun. 7, 2016, which is a U.S. National Phaseapplication of International Patent Application No. PCT/US2014/069373,filed on Dec. 9, 2014, which claims the benefit of and priority to U.S.Provisional Application No. 61/914,275, filed on Dec. 10, 2013; each ofthe aforementioned disclosures is incorporated herein by reference inits entirety.

BACKGROUND

Embodiments of the present disclosure relate generally to biological orchemical analysis and more particularly to systems and methods usingdetection devices for biological or chemical analysis.

Various protocols in biological or chemical research involve performinga large number of controlled reactions on local support surfaces orwithin predefined reaction chambers. The designated reactions may thenbe observed or detected and subsequent analysis may help identify orreveal properties of chemicals involved in the reaction. For example, insome multiplex assays, an unknown analyte having an identifiable label(e.g., fluorescent label) may be exposed to thousands of known probesunder controlled conditions. Each known probe may be deposited into acorresponding well of a microplate. Observing any chemical reactionsthat occur between the known probes and the unknown analyte within thewells may help identify or reveal properties of the analyte. Otherexamples of such protocols include known DNA sequencing processes, suchas sequencing-by-synthesis (SBS) or cyclic-array sequencing.

In some conventional fluorescent-detection protocols, an optical systemis used to direct an excitation light onto fluorescently-labeledanalytes and to also detect the fluorescent signals that may emit fromthe analytes. However, such optical systems can be relatively expensiveand require a larger benchtop footprint. For example, the optical systemmay include an arrangement of lenses, filters, and light sources. Inother proposed detection systems, the controlled reactions occurimmediately over a solid-state imager (e.g., charged-coupled device(CCD) or a complementary metal-oxide-semiconductor (CMOS) detector) thatdoes not require a large optical assembly to detect the fluorescentemissions.

However, the proposed solid-state imaging systems may have somelimitations. For example, it may be challenging to distinguish thefluorescent emissions from the excitation light when the excitationlight is also directed toward the light sensors of the solid-stateimager. In addition, fluidicly delivering reagents to analytes that arelocated on an electronic device and in a controlled manner may presentadditional challenges. As another example, fluorescent emissions aresubstantially isotropic. As the density of the analytes on thesolid-state imager increases, it becomes increasingly challenging tomanage or account for unwanted light emissions from adjacent analytes(e.g., crosstalk).

BRIEF DESCRIPTION

In an embodiment, a biosensor is provided that includes a flow cell anda detection device having the flow cell coupled thereto. The flow celland the detection device form a flow channel that is configured to havebiological or chemical substances therein that generate light emissionsin response to an excitation light. The detection device includes adevice base having a sensor array of light sensors and a guide array oflight guides. The light guides have input regions that are configured toreceive the excitation light and the light emissions from the flowchannel. The light guides extend into the device base from the inputregions toward corresponding light sensors and have a filter materialthat is configured to filter the excitation light and permit the lightemissions to propagate toward the corresponding light sensors. Thedevice base includes device circuitry electrically coupled to the lightsensors and configured to transmit data signals based on photonsdetected by the light sensors. The detection device also includes ashield layer that extends between the flow channel and the device base.The shield layer has apertures that are positioned relative to the inputregions of corresponding light guides such that the light emissionspropagate through the apertures into the corresponding input regions.The shield layer extends between adjacent apertures and is configured toblock the excitation light and the light emissions incident on theshield layer between the adjacent apertures.

In an embodiment, a biosensor is provided that includes a flow cell anda detection device having the flow cell coupled thereto. The flow celland the detection device form a flow channel that is configured to havebiological or chemical substances therein that generate light emissionsin response to an excitation light. The detection device may include adevice base having a sensor array of light sensors and a guide array oflight guides. The light guides are configured to receive the excitationlight and the light emissions from the flow channel. Each of the lightguides extends into the device base along a central longitudinal axisfrom an input region of the light guide toward a corresponding lightsensor of the sensor array. The light guides include a filter materialthat is configured to filter the excitation light and permit the lightemissions to propagate therethrough toward the corresponding lightsensors. The device base includes device circuitry that is electricallycoupled to the light sensors and configured to transmit data signalsbased on photons detected by the light sensors. The device base includesperipheral crosstalk shields located therein that surround correspondinglight guides of the guide array. The crosstalk shields at leastpartially surround the corresponding light guides about the respectivelongitudinal axis to reduce optical crosstalk between adjacent lightsensors.

In an embodiment, a method of manufacturing a biosensor is provided. Themethod includes providing a device base having a sensor array of lightsensors and device circuitry that is electrically coupled to the lightsensors and configured to transmit data signals based on photonsdetected by the light sensors. The device base has an outer surface. Themethod also includes applying a shield layer to the outer surface of thedevice base and forming apertures through the shield layer. The methodalso includes forming guide cavities that extend from correspondingapertures toward a corresponding light sensor of the sensor array anddepositing filter material within the guide cavities. A portion of thefilter material extends along the shield layer. The method also includescuring the filter material and removing the filter material from theshield layer. The filter material within the guide cavities forms lightguides. The method also includes applying a passivation layer to theshield layer such that the passivation layer extends directly along theshield layer and across the apertures.

In an embodiment, a biosensor is provided that includes a device basehaving a sensor array of light sensors and a guide array of lightguides. The device base has an outer surface. The light guides haveinput regions that are configured to receive excitation light and lightemissions generated by biological or chemical substances proximate tothe outer surface. The light guides extend into the device base from theinput regions toward corresponding light sensors and have a filtermaterial that is configured to filter the excitation light and permitthe light emissions to propagate toward the corresponding light sensors.The device base includes device circuitry electrically coupled to thelight sensors and configured to transmit data signals based on photonsdetected by the light sensors. The biosensor also includes a shieldlayer that extends along the outer surface of the device base. Theshield layer has apertures that are positioned relative to the inputregions of corresponding light guides such that the light emissionspropagate through the apertures into the corresponding input regions.The shield layer extends between adjacent apertures and is configured toblock the excitation light and the light emissions incident on theshield layer between the adjacent apertures.

In an embodiment, a biosensor is provided that includes a device basehaving a sensor array of light sensors and a guide array of lightguides. The device base has an outer surface. The light guides areconfigured to receive excitation light and light emissions generated bybiological or chemical substances proximate to the outer surface. Eachof the light guides extends into the device base along a centrallongitudinal axis from an input region of the light guide toward acorresponding light sensor of the sensor array. The light guide includesa filter material that is configured to filter the excitation light andpermit the light emissions to propagate therethrough towardcorresponding light sensors. The device base includes device circuitryelectrically coupled to the light sensors and configured to transmitdata signals based on photons detected by the light sensors. The devicebase includes peripheral crosstalk shields located therein that surroundcorresponding light guides of the guide array. The crosstalk shields atleast partially surrounding the corresponding light guides about therespective longitudinal axis to at least one of block or reflect errantlight rays to reduce optical crosstalk between adjacent light sensors.

While multiple embodiments are described, still other embodiments of thedescribed subject matter will become apparent to those skilled in theart from the following detailed description and drawings, which show anddescribe illustrative embodiments of disclosed inventive subject matter.As will be realized, the inventive subject matter is capable ofmodifications in various aspects, all without departing from the spiritand scope of the described subject matter. Accordingly, the drawings anddetailed description are to be regarded as illustrative in nature andnot restrictive.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a block diagram of an exemplary system for biological orchemical analysis formed in accordance with one embodiment.

FIG. 2 is a block diagram of an exemplary system controller that may beused in the system of FIG. 1.

FIG. 3 is a block diagram of an exemplary workstation for biological orchemical analysis in accordance with one embodiment.

FIG. 4 is a perspective view of an exemplary workstation and anexemplary cartridge in accordance with one embodiment.

FIG. 5 is a front view of an exemplary rack assembly that includes aplurality of the workstations of FIG. 4.

FIG. 6 illustrates internal components of an exemplary cartridge.

FIG. 7 illustrates a cross-section of a biosensor formed in accordancewith one embodiment.

FIG. 8 is an enlarged portion of the cross-section of FIG. 7illustrating the biosensor in greater detail.

FIG. 9 is another enlarged portion of the cross-section of FIG. 7illustrating the biosensor in greater detail.

FIG. 10 is a schematic cross-section of a detection device formed inaccordance with another embodiment.

FIG. 11 is a flowchart illustrating a method of manufacturing abiosensor in accordance with an embodiment.

FIGS. 12A and 12B illustrate different stages of manufacturing thebiosensor of FIG. 11.

DETAILED DESCRIPTION

Embodiments described herein may be used in various biological orchemical processes and systems for academic or commercial analysis. Morespecifically, embodiments described herein may be used in variousprocesses and systems where it is desired to detect an event, property,quality, or characteristic that is indicative of a designated reaction.For example, embodiments described herein include cartridges,biosensors, and their components as well as bioassay systems thatoperate with cartridges and biosensors. In particular embodiments, thecartridges and biosensors include a flow cell and one or more lightsensors that are coupled together in a substantially unitary structure.

The bioassay systems may be configured to perform a plurality ofdesignated reactions that may be detected individually or collectively.The biosensors and bioassay systems may be configured to performnumerous cycles in which the plurality of designated reactions occurs inparallel. For example, the bioassay systems may be used to sequence adense array of DNA features through iterative cycles of enzymaticmanipulation and image acquisition. As such, the cartridges andbiosensors may include one or more microfluidic channels that deliverreagents or other reaction components to a reaction site. In someembodiments, the reaction sites are randomly distributed across asubstantially planer surface. For example, the reaction sites may havean uneven distribution in which some reaction sites are located closerto each other than other reaction sites. In other embodiments, thereaction sites are patterned across a substantially planer surface in apredetermined manner. Each of the reaction sites may be associated withone or more light sensors that detect light from the associated reactionsite. Yet in other embodiments, the reaction sites are located inreaction chambers that compartmentalize the designated reactionstherein.

The following detailed description of certain embodiments will be betterunderstood when read in conjunction with the appended drawings. To theextent that the figures illustrate diagrams of the functional blocks ofvarious embodiments, the functional blocks are not necessarilyindicative of the division between hardware circuitry. Thus, forexample, one or more of the functional blocks (e.g., processors ormemories) may be implemented in a single piece of hardware (e.g., ageneral purpose signal processor or random access memory, hard disk, orthe like). Similarly, the programs may be stand alone programs, may beincorporated as subroutines in an operating system, may be functions inan installed software package, and the like. It should be understoodthat the various embodiments are not limited to the arrangements andinstrumentality shown in the drawings.

As used herein, an element or step recited in the singular and proceededwith the word “a” or “an” should be understood as not excluding pluralof said elements or steps, unless such exclusion is explicitly stated.Furthermore, references to “one embodiment” are not intended to beinterpreted as excluding the existence of additional embodiments thatalso incorporate the recited features. Moreover, unless explicitlystated to the contrary, embodiments “comprising” or “having” an elementor a plurality of elements having a particular property may includeadditional elements whether or not they have that property.

As used herein, a “designated reaction” includes a change in at leastone of a chemical, electrical, physical, or optical property (orquality) of an analyte-of-interest. In particular embodiments, thedesignated reaction is a positive binding event (e.g., incorporation ofa fluorescently labeled biomolecule with the analyte-of-interest). Moregenerally, the designated reaction may be a chemical transformation,chemical change, or chemical interaction. The designated reaction mayalso be a change in electrical properties. For example, the designatedreaction may be a change in ion concentration within a solution.Exemplary reactions include, but are not limited to, chemical reactionssuch as reduction, oxidation, addition, elimination, rearrangement,esterification, amidation, etherification, cyclization, or substitution;binding interactions in which a first chemical binds to a secondchemical; dissociation reactions in which two or more chemicals detachfrom each other; fluorescence; luminescence; bioluminescence;chemiluminescence; and biological reactions, such as nucleic acidreplication, nucleic acid amplification, nucleic acid hybridization,nucleic acid ligation, phosphorylation, enzymatic catalysis, receptorbinding, or ligand binding. The designated reaction can also be additionor elimination of a proton, for example, detectable as a change in pH ofa surrounding solution or environment. An additional designated reactioncan be detecting the flow of ions across a membrane (e.g., natural orsynthetic bilayer membrane), for example as ions flow through a membranethe current is disrupted and the disruption can be detected.

In particular embodiments, the designated reaction includes theincorporation of a fluorescently-labeled molecule to an analyte. Theanalyte may be an oligonucleotide and the fluorescently-labeled moleculemay be a nucleotide. The designated reaction may be detected when anexcitation light is directed toward the oligonucleotide having thelabeled nucleotide, and the fluorophore emits a detectable fluorescentsignal. In alternative embodiments, the detected fluorescence is aresult of chemiluminescence or bioluminescence. A designated reactionmay also increase fluorescence (or Förster) resonance energy transfer(FRET), for example, by bringing a donor fluorophore in proximity to anacceptor fluorophore, decrease FRET by separating donor and acceptorfluorophores, increase fluorescence by separating a quencher from afluorophore or decrease fluorescence by co-locating a quencher andfluorophore.

As used herein, a “reaction component” or “reactant” includes anysubstance that may be used to obtain a designated reaction. For example,reaction components include reagents, enzymes, samples, otherbiomolecules, and buffer solutions. The reaction components aretypically delivered to a reaction site in a solution and/or immobilizedat a reaction site. The reaction components may interact directly orindirectly with another substance, such as the analyte-of-interest.

As used herein, the term “reaction site” is a localized region where adesignated reaction may occur. A reaction site may include supportsurfaces of a substrate where a substance may be immobilized thereon.For example, a reaction site may include a substantially planar surfacein a channel of a flow cell that has a colony of nucleic acids thereon.Typically, but not always, the nucleic acids in the colony have the samesequence, being for example, clonal copies of a single stranded ordouble stranded template. However, in some embodiments a reaction sitemay contain only a single nucleic acid molecule, for example, in asingle stranded or double stranded form. Furthermore, a plurality ofreaction sites may be randomly distributed along the support surface orarranged in a predetermined manner (e.g., side-by-side in a matrix, suchas in microarrays). A reaction site can also include a reaction chamberthat at least partially defines a spatial region or volume configured tocompartmentalize the designated reaction. As used herein, the term“reaction chamber” includes a spatial region that is in fluidcommunication with a flow channel. The reaction chamber may be at leastpartially separated from the surrounding environment or other spatialregions. For example, a plurality of reaction chambers may be separatedfrom each other by shared walls. As a more specific example, thereaction chamber may include a cavity defined by interior surfaces of awell and have an opening or aperture so that the cavity may be in fluidcommunication with a flow channel. Biosensors including such reactionchambers are described in greater detail in international applicationno. PCT/US2011/057111, filed on Oct. 20, 2011, which is incorporatedherein by reference in its entirety.

In some embodiments, the reaction chambers are sized and shaped relativeto solids (including semi-solids) so that the solids may be inserted,fully or partially, therein. For example, the reaction chamber may besized and shaped to accommodate only one capture bead. The capture beadmay have clonally amplified DNA or other substances thereon.Alternatively, the reaction chamber may be sized and shaped to receivean approximate number of beads or solid substrates. As another example,the reaction chambers may also be filled with a porous gel or substancethat is configured to control diffusion or filter fluids that may flowinto the reaction chamber.

In some embodiments, light sensors (e.g., photodiodes) are associatedwith corresponding reaction sites. A light sensor that is associatedwith a reaction site is configured to detect light emissions from theassociated reaction site when a designated reaction has occurred at theassociated reaction site. In some cases, a plurality of light sensors(e.g. several pixels of a camera device) may be associated with a singlereaction site. In other cases, a single light sensor (e.g. a singlepixel) may be associated with a single reaction site or with a group ofreaction sites. The light sensor, the reaction site, and other featuresof the biosensor may be configured so that at least some of the light isdirectly detected by the light sensor without being reflected.

As used herein, the term “adjacent” when used with respect to tworeaction sites means no other reaction site is located between the tworeaction sites. The term “adjacent” may have a similar meaning when usedwith respect to adjacent detection paths and adjacent light sensors(e.g., adjacent light sensors have no other light sensor therebetween).In some cases, a reaction site may not be adjacent to another reactionsite, but may still be within an immediate vicinity of the otherreaction site. A first reaction site may be in the immediate vicinity ofa second reaction site when fluorescent emission signals from the firstreaction site are detected by the light sensor associated with thesecond reaction site. More specifically, a first reaction site may be inthe immediate vicinity of a second reaction site when the light sensorassociated with the second reaction site detects, for example crosstalkfrom the first reaction site. Adjacent reaction sites can be contiguoussuch that they abut each other or the adjacent sites can benon-contiguous having an intervening space between.

As used herein, a “substance” includes items or solids, such as capturebeads, as well as biological or chemical substances. As used herein, a“biological or chemical substance” includes biomolecules,samples-of-interest, analytes-of-interest, and other chemicalcompound(s). A biological or chemical substance may be used to detect,identify, or analyze other chemical compound(s), or function asintermediaries to study or analyze other chemical compound(s). Inparticular embodiments, the biological or chemical substances include abiomolecule. As used herein, a “biomolecule” includes at least one of abiopolymer, nucleoside, nucleic acid, polynucleotide, oligonucleotide,protein, enzyme, polypeptide, antibody, antigen, ligand, receptor,polysaccharide, carbohydrate, polyphosphate, cell, tissue, organism, orfragment thereof or any other biologically active chemical compound(s)such as analogs or mimetics of the aforementioned species.

In a further example, a biological or chemical substance or abiomolecule includes an enzyme or reagent used in a coupled reaction todetect the product of another reaction such as an enzyme or reagent usedto detect pyrophosphate in a pyrosequencing reaction. Enzymes andreagents useful for pyrophosphate detection are described, for example,in U.S. Patent Publication No. 2005/0244870 A1, which is incorporatedherein in its entirety.

Biomolecules, samples, and biological or chemical substances may benaturally occurring or synthetic and may be suspended in a solution ormixture within a spatial region. Biomolecules, samples, and biologicalor chemical substances may also be bound to a solid phase or gelmaterial. Biomolecules, samples, and biological or chemical substancesmay also include a pharmaceutical composition. In some cases,biomolecules, samples, and biological or chemical substances of interestmay be referred to as targets, probes, or analytes.

As used herein, a “biosensor” includes a structure having a plurality ofreaction sites that is configured to detect designated reactions thatoccur at or proximate to the reaction sites. A biosensor may include asolid-state imaging device (e.g., CCD or CMOS imager) and, optionally, aflow cell mounted thereto. The flow cell may include at least one flowchannel that is in fluid communication with the reaction sites. As onespecific example, the biosensor is configured to fluidicly andelectrically couple to a bioassay system. The bioassay system maydeliver reactants to the reaction sites according to a predeterminedprotocol (e.g., sequencing-by-synthesis) and perform a plurality ofimaging events. For example, the bioassay system may direct solutions toflow along the reaction sites. At least one of the solutions may includefour types of nucleotides having the same or different fluorescentlabels. The nucleotides may bind to corresponding oligonucleotideslocated at the reaction sites. The bioassay system may then illuminatethe reaction sites using an excitation light source (e.g., solid-statelight sources, such as light-emitting diodes or LEDs). The excitationlight may have a predetermined wavelength or wavelengths, including arange of wavelengths. The excited fluorescent labels provide emissionsignals that may be detected by the light sensors.

In alternative embodiments, the biosensor may include electrodes orother types of sensors configured to detect other identifiableproperties. For example, the sensors may be configured to detect achange in ion concentration. In another example, the sensors may beconfigured to detect the ion current flow across a membrane

As used herein, a “cartridge” includes a structure that is configured tohold a biosensor. In some embodiments, the cartridge may includeadditional features, such as the light source (e.g., LEDs) that areconfigured to provide excitation light to the reactions sites of thebiosensor. The cartridge may also include a fluidic storage system(e.g., storage for reagents, sample, and buffer) and a fluidic controlsystem (e.g., pumps, valves, and the like) for fluidically transportingreaction components, sample, and the like to the reaction sites. Forexample, after the biosensor is prepared or manufactured, the biosensormay be coupled to a housing or container of the cartridge. In someembodiments, the biosensors and the cartridges may be self-contained,disposable units. However, other embodiments may include an assemblywith removable parts that allow a user to access an interior of thebiosensor or cartridge for maintenance or replacement of components orsamples. The biosensor and the cartridge may be removably coupled orengaged to larger bioassay systems, such as a sequencing system, thatconducts controlled reactions therein.

As used herein, when the terms “removably” and “coupled” (or “engaged”)are used together to describe a relationship between the biosensor (orcartridge) and a system receptacle or interface of a bioassay system,the term is intended to mean that a connection between the biosensor (orcartridge) and the system receptacle is readily separable withoutdestroying or damaging the system receptacle and/or the biosensor (orcartridge). Components are readily separable when the components may beseparated from each other without undue effort or a significant amountof time spent in separating the components. For example, the biosensor(or cartridge) may be removably coupled or engaged to the systemreceptacle in an electrical manner such that the mating contacts of thebioassay system are not destroyed or damaged. The biosensor (orcartridge) may also be removably coupled or engaged to the systemreceptacle in a mechanical manner such that the features that hold thebiosensor (or cartridge) are not destroyed or damaged. The biosensor (orcartridge) may also be removably coupled or engaged to the systemreceptacle in a fluidic manner such that the ports of the systemreceptacle are not destroyed or damaged. The system receptacle or acomponent is not considered to be destroyed or damaged if, for example,only a simple adjustment to the component (e.g., realignment) or asimple replacement (e.g., replacing a nozzle) is required.

As used herein, the term “fluid communication” or “fluidicly coupled”refers to two spatial regions being connected together such that aliquid or gas may flow between the two spatial regions. For example, amicrofluidic channel may be in fluid communication with a reactionchamber such that a fluid may flow freely into the reaction chamber fromthe microfluidic channel. The terms “in fluid communication” or“fluidicly coupled” allow for two spatial regions being in fluidcommunication through one or more valves, restrictors, or other fluidiccomponents that are configured to control or regulate a flow of fluidthrough a system.

As used herein, the term “immobilized,” when used with respect to abiomolecule or biological or chemical substance, includes substantiallyattaching the biomolecule or biological or chemical substance at amolecular level to a surface. For example, a biomolecule or biologicalor chemical substance may be immobilized to a surface of the substratematerial using adsorption techniques including non-covalent interactions(e.g., electrostatic forces, van der Waals, and dehydration ofhydrophobic interfaces) and covalent binding techniques where functionalgroups or linkers facilitate attaching the biomolecules to the surface.Immobilizing biomolecules or biological or chemical substances to asurface of a substrate material may be based upon the properties of thesubstrate surface, the liquid medium carrying the biomolecule orbiological or chemical substance, and the properties of the biomoleculesor biological or chemical substances themselves. In some cases, asubstrate surface may be functionalized (e.g., chemically or physicallymodified) to facilitate immobilizing the biomolecules (or biological orchemical substances) to the substrate surface. The substrate surface maybe first modified to have functional groups bound to the surface. Thefunctional groups may then bind to biomolecules or biological orchemical substances to immobilize them thereon. A substance can beimmobilized to a surface via a gel, for example, as described in USPatent Publ. No. US 2011/0059865 A1, which is incorporated herein byreference.

In some embodiments, nucleic acids can be attached to a surface andamplified using bridge amplification. Useful bridge amplificationmethods are described, for example, in U.S. Pat. No. 5,641,658; WO07/010251, U.S. Pat. No. 6,090,592; U.S. Patent Publ. No. 2002/0055100A1; U.S. Pat. No. 7,115,400; U.S. Patent Publ. No. 2004/0096853 A1; U.S.Patent Publ. No. 2004/0002090 A1; U.S. Patent Publ. No. 2007/0128624 A1;and U.S. Patent Publ. No. 2008/0009420 A1, each of which is incorporatedherein in its entirety. Another useful method for amplifying nucleicacids on a surface is rolling circle amplification (RCA), for example,using methods set forth in further detail below. In some embodiments,the nucleic acids can be attached to a surface and amplified using oneor more primer pairs. For example, one of the primers can be in solutionand the other primer can be immobilized on the surface (e.g.,5′-attached). By way of example, a nucleic acid molecule can hybridizeto one of the primers on the surface followed by extension of theimmobilized primer to produce a first copy of the nucleic acid. Theprimer in solution then hybridizes to the first copy of the nucleic acidwhich can be extended using the first copy of the nucleic acid as atemplate. Optionally, after the first copy of the nucleic acid isproduced, the original nucleic acid molecule can hybridize to a secondimmobilized primer on the surface and can be extended at the same timeor after the primer in solution is extended. In any embodiment, repeatedrounds of extension (e.g., amplification) using the immobilized primerand primer in solution provide multiple copies of the nucleic acid.

In particular embodiments, the assay protocols executed by the systemsand methods described herein include the use of natural nucleotides andalso enzymes that are configured to interact with the naturalnucleotides. Natural nucleotides include, for example, ribonucleotidesor deoxyribonucleotides. Natural nucleotides can be in the mono-, di-,or tri-phosphate form and can have a base selected from adenine (A),Thymine (T), uracil (U), guanine (G) or cytosine (C). It will beunderstood however that non-natural nucleotides, modified nucleotides oranalogs of the aforementioned nucleotides can be used. Some examples ofuseful non-natural nucleotides are set forth below in regard toreversible terminator-based sequencing by synthesis methods.

In embodiments that include reaction chambers, items or solid substances(including semi-solid substances) may be disposed within the reactionchambers. When disposed, the item or solid may be physically held orimmobilized within the reaction chamber through an interference fit,adhesion, or entrapment. Exemplary items or solids that may be disposedwithin the reaction chambers include polymer beads, pellets, agarosegel, powders, quantum dots, or other solids that may be compressedand/or held within the reaction chamber. In particular embodiments, anucleic acid superstructure, such as a DNA ball, can be disposed in orat a reaction chamber, for example, by attachment to an interior surfaceof the reaction chamber or by residence in a liquid within the reactionchamber. A DNA ball or other nucleic acid superstructure can bepreformed and then disposed in or at the reaction chamber.Alternatively, a DNA ball can be synthesized in situ at the reactionchamber. A DNA ball can be synthesized by rolling circle amplificationto produce a concatamer of a particular nucleic acid sequence and theconcatamer can be treated with conditions that form a relatively compactball. DNA balls and methods for their synthesis are described, forexample in, U.S. Patent Publ. Nos. 2008/0242560 A1 or 2008/0234136 A1,each of which is incorporated herein in its entirety. A substance thatis held or disposed in a reaction chamber can be in a solid, liquid, orgaseous state.

FIG. 1 is a block diagram of an exemplary bioassay system 100 forbiological or chemical analysis formed in accordance with oneembodiment. The term “bioassay” is not intended to be limiting as thebioassay system 100 may operate to obtain any information or data thatrelates to at least one of a biological or chemical substance. In someembodiments, the bioassay system 100 is a workstation that may besimilar to a bench-top device or desktop computer. For example, amajority (or all) of the systems and components for conducting thedesignated reactions can be within a common housing 116.

In particular embodiments, the bioassay system 100 is a nucleic acidsequencing system (or sequencer) configured for various applications,including but not limited to de novo sequencing, resequencing of wholegenomes or target genomic regions, and metagenomics. The sequencer mayalso be used for DNA or RNA analysis. In some embodiments, the bioassaysystem 100 may also be configured to generate reaction sites in abiosensor. For example, the bioassay system 100 may be configured toreceive a sample and generate surface attached clusters of clonallyamplified nucleic acids derived from the sample. Each cluster mayconstitute or be part of a reaction site in the biosensor.

The exemplary bioassay system 100 may include a system receptacle orinterface 112 that is configured to interact with a biosensor 102 toperform designated reactions within the biosensor 102. In the followingdescription with respect to FIG. 1, the biosensor 102 is loaded into thesystem receptacle 112. However, it is understood that a cartridge thatincludes the biosensor 102 may be inserted into the system receptacle112 and in some states the cartridge can be removed temporarily orpermanently. As described above, the cartridge may include, among otherthings, fluidic control and fluidic storage components.

In particular embodiments, the bioassay system 100 is configured toperform a large number of parallel reactions within the biosensor 102.The biosensor 102 includes one or more reaction sites where designatedreactions can occur. The reaction sites may be, for example, immobilizedto a solid surface of the biosensor or immobilized to beads (or othermovable substrates) that are located within corresponding reactionchambers of the biosensor. The reaction sites can include, for example,clusters of clonally amplified nucleic acids. The biosensor 102 mayinclude a solid-state imaging device (e.g., CCD or CMOS imager) and aflow cell mounted thereto. The flow cell may include one or more flowchannels that receive a solution from the bioassay system 100 and directthe solution toward the reaction sites. Optionally, the biosensor 102can be configured to engage a thermal element for transferring thermalenergy into or out of the flow channel.

The bioassay system 100 may include various components, assemblies, andsystems (or sub-systems) that interact with each other to perform apredetermined method or assay protocol for biological or chemicalanalysis. For example, the bioassay system 100 includes a systemcontroller 104 that may communicate with the various components,assemblies, and sub-systems of the bioassay system 100 and also thebiosensor 102. For example, in addition to the system receptacle 112,the bioassay system 100 may also include a fluidic control system 106 tocontrol the flow of fluid throughout a fluid network of the bioassaysystem 100 and the biosensor 102; a fluid storage system 108 that isconfigured to hold all fluids (e.g., gas or liquids) that may be used bythe bioassay system; a temperature control system 110 that may regulatethe temperature of the fluid in the fluid network, the fluid storagesystem 108, and/or the biosensor 102; and an illumination system 111that is configured to illuminate the biosensor 102. As described above,if a cartridge having the biosensor 102 is loaded into the systemreceptacle 112, the cartridge may also include fluidic control andfluidic storage components.

Also shown, the bioassay system 100 may include a user interface 114that interacts with the user. For example, the user interface 114 mayinclude a display 113 to display or request information from a user anda user input device 115 to receive user inputs. In some embodiments, thedisplay 113 and the user input device 115 are the same device. Forexample, the user interface 114 may include a touch-sensitive displayconfigured to detect the presence of an individual's touch and alsoidentify a location of the touch on the display. However, other userinput devices 115 may be used, such as a mouse, touchpad, keyboard,keypad, handheld scanner, voice-recognition system, motion-recognitionsystem, and the like. As will be discussed in greater detail below, thebioassay system 100 may communicate with various components, includingthe biosensor 102 (e.g. in the form of a cartridge), to perform thedesignated reactions. The bioassay system 100 may also be configured toanalyze data obtained from the biosensor to provide a user with desiredinformation.

The system controller 104 may include any processor-based ormicroprocessor-based system, including systems using microcontrollers,reduced instruction set computers (RISC), application specificintegrated circuits (ASICs), field programmable gate array (FPGAs),logic circuits, and any other circuit or processor capable of executingfunctions described herein. The above examples are exemplary only, andare thus not intended to limit in any way the definition and/or meaningof the term system controller. In the exemplary embodiment, the systemcontroller 104 executes a set of instructions that are stored in one ormore storage elements, memories, or modules in order to at least one ofobtain and analyze detection data. Storage elements may be in the formof information sources or physical memory elements within the bioassaysystem 100.

The set of instructions may include various commands that instruct thebioassay system 100 or biosensor 102 to perform specific operations suchas the methods and processes of the various embodiments describedherein. The set of instructions may be in the form of a softwareprogram, which may form part of a tangible, non-transitory computerreadable medium or media. As used herein, the terms “software” and“firmware” are interchangeable, and include any computer program storedin memory for execution by a computer, including RAM memory, ROM memory,EPROM memory, EEPROM memory, and non-volatile RAM (NVRAM) memory. Theabove memory types are exemplary only, and are thus not limiting as tothe types of memory usable for storage of a computer program.

The software may be in various forms such as system software orapplication software. Further, the software may be in the form of acollection of separate programs, or a program module within a largerprogram or a portion of a program module. The software also may includemodular programming in the form of object-oriented programming. Afterobtaining the detection data, the detection data may be automaticallyprocessed by the bioassay system 100, processed in response to userinputs, or processed in response to a request made by another processingmachine (e.g., a remote request through a communication link).

The system controller 104 may be connected to the biosensor 102 and theother components of the bioassay system 100 via communication links. Thesystem controller 104 may also be communicatively connected to off-sitesystems or servers. The communication links may be hardwired orwireless. The system controller 104 may receive user inputs or commands,from the user interface 114 and the user input device 115.

The fluidic control system 106 includes a fluid network and isconfigured to direct and regulate the flow of one or more fluids throughthe fluid network. The fluid network may be in fluid communication withthe biosensor 102 and the fluid storage system 108. For example, selectfluids may be drawn from the fluid storage system 108 and directed tothe biosensor 102 in a controlled manner, or the fluids may be drawnfrom the biosensor 102 and directed toward, for example, a wastereservoir in the fluid storage system 108. Although not shown, thefluidic control system 106 may include flow sensors that detect a flowrate or pressure of the fluids within the fluid network. The sensors maycommunicate with the system controller 104.

The temperature control system 110 is configured to regulate thetemperature of fluids at different regions of the fluid network, thefluid storage system 108, and/or the biosensor 102. For example, thetemperature control system 110 may include a thermocycler thatinterfaces with the biosensor 102 and controls the temperature of thefluid that flows along the reaction sites in the biosensor 102. Thetemperature control system 110 may also regulate the temperature ofsolid elements or components of the bioassay system 100 or the biosensor102. Although not shown, the temperature control system 110 may includesensors to detect the temperature of the fluid or other components. Thesensors may communicate with the system controller 104.

The fluid storage system 108 is in fluid communication with thebiosensor 102 and may store various reaction components or reactantsthat are used to conduct the designated reactions therein. The fluidstorage system 108 may also store fluids for washing or cleaning thefluid network and biosensor 102 and for diluting the reactants. Forexample, the fluid storage system 108 may include various reservoirs tostore samples, reagents, enzymes, other biomolecules, buffer solutions,aqueous, and non-polar solutions, and the like. Furthermore, the fluidstorage system 108 may also include waste reservoirs for receiving wasteproducts from the biosensor 102. In embodiments that include acartridge, the cartridge may include one or more of a fluid storagesystem, fluidic control system or temperature control system.Accordingly, one or more of the components set forth herein as relatingto those systems can be contained within a cartridge housing. Forexample, a cartridge can have various reservoirs to store samples,reagents, enzymes, other biomolecules, buffer solutions, aqueous, andnon-polar solutions, waste, and the like. As such, one or more of afluid storage system, fluidic control system or temperature controlsystem can be removably engaged with a bioassay system via a cartridgeor other biosensor.

The illumination system 111 may include a light source (e.g., one ormore LEDs) and a plurality of optical components to illuminate thebiosensor. Examples of light sources may include lasers, arc lamps,LEDs, or laser diodes. The optical components may be, for example,reflectors, dichroics, beam splitters, collimators, lenses, filters,wedges, prisms, mirrors, detectors, and the like. In embodiments thatuse an illumination system, the illumination system 111 may beconfigured to direct an excitation light to reaction sites. As oneexample, fluorophores may be excited by green wavelengths of light, assuch the wavelength of the excitation light may be approximately 532 nm.

The system receptacle or interface 112 is configured to engage thebiosensor 102 in at least one of a mechanical, electrical, and fluidicmanner. The system receptacle 112 may hold the biosensor 102 in adesired orientation to facilitate the flow of fluid through thebiosensor 102. The system receptacle 112 may also include electricalcontacts that are configured to engage the biosensor 102 so that thebioassay system 100 may communicate with the biosensor 102 and/orprovide power to the biosensor 102. Furthermore, the system receptacle112 may include fluidic ports (e.g., nozzles) that are configured toengage the biosensor 102. In some embodiments, the biosensor 102 isremovably coupled to the system receptacle 112 in a mechanical manner,in an electrical manner, and also in a fluidic manner.

In addition, the bioassay system 100 may communicate remotely with othersystems or networks or with other bioassay systems 100. Detection dataobtained by the bioassay system(s) 100 may be stored in a remotedatabase.

FIG. 2 is a block diagram of the system controller 104 in the exemplaryembodiment. In one embodiment, the system controller 104 includes one ormore processors or modules that can communicate with one another. Eachof the processors or modules may include an algorithm (e.g.,instructions stored on a tangible and/or non-transitory computerreadable storage medium) or sub-algorithms to perform particularprocesses. The system controller 104 is illustrated conceptually as acollection of modules, but may be implemented utilizing any combinationof dedicated hardware boards, DSPs, processors, etc. Alternatively, thesystem controller 104 may be implemented utilizing an off-the-shelf PCwith a single processor or multiple processors, with the functionaloperations distributed between the processors. As a further option, themodules described below may be implemented utilizing a hybridconfiguration in which certain modular functions are performed utilizingdedicated hardware, while the remaining modular functions are performedutilizing an off-the-shelf PC and the like. The modules also may beimplemented as software modules within a processing unit.

During operation, a communication link 120 may transmit information(e.g. commands) to or receive information (e.g. data) from the biosensor102 (FIG. 1) and/or the sub-systems 106, 108, 110 (FIG. 1). Acommunication link 122 may receive user input from the user interface114 (FIG. 1) and transmit data or information to the user interface 114.Data from the biosensor 102 or sub-systems 106, 108, 110 may beprocessed by the system controller 104 in real-time during a bioassaysession. Additionally or alternatively, data may be stored temporarilyin a system memory during a bioassay session and processed in slowerthan real-time or off-line operation.

As shown in FIG. 2, the system controller 104 may include a plurality ofmodules 131-139 that communicate with a main control module 130. Themain control module 130 may communicate with the user interface 114(FIG. 1). Although the modules 131-139 are shown as communicatingdirectly with the main control module 130, the modules 131-139 may alsocommunicate directly with each other, the user interface 114, and thebiosensor 102. Also, the modules 131-139 may communicate with the maincontrol module 130 through the other modules.

The plurality of modules 131-139 include system modules 131-133, 139that communicate with the sub-systems 106, 108, 110, and 111,respectively. The fluidic control module 131 may communicate with thefluidic control system 106 to control the valves and flow sensors of thefluid network for controlling the flow of one or more fluids through thefluid network. The fluid storage module 132 may notify the user whenfluids are low or when the waste reservoir is at or near capacity. Thefluid storage module 132 may also communicate with the temperaturecontrol module 133 so that the fluids may be stored at a desiredtemperature. The illumination module 139 may communicate with theillumination system 109 to illuminate the reaction sites at designatedtimes during a protocol, such as after the designated reactions (e.g.,binding events) have occurred.

The plurality of modules 131-139 may also include a device module 134that communicates with the biosensor 102 and an identification module135 that determines identification information relating to the biosensor102. The device module 134 may, for example, communicate with the systemreceptacle 112 to confirm that the biosensor has established anelectrical and fluidic connection with the bioassay system 100. Theidentification module 135 may receive signals that identify thebiosensor 102. The identification module 135 may use the identity of thebiosensor 102 to provide other information to the user. For example, theidentification module 135 may determine and then display a lot number, adate of manufacture, or a protocol that is recommended to be run withthe biosensor 102.

The plurality of modules 131-139 may also include a detection dataanalysis module 138 that receives and analyzes the signal data (e.g.,image data) from the biosensor 102. The signal data may be stored forsubsequent analysis or may be transmitted to the user interface 114 todisplay desired information to the user. In some embodiments, the signaldata may be processed by the solid-state imager (e.g., CMOS imagesensor) before the detection data analysis module 138 receives thesignal data.

Protocol modules 136 and 137 communicate with the main control module130 to control the operation of the sub-systems 106, 108, and 110 whenconducting predetermined assay protocols. The protocol modules 136 and137 may include sets of instructions for instructing the bioassay system100 to perform specific operations pursuant to predetermined protocols.As shown, the protocol module may be a sequencing-by-synthesis (SBS)module 136 that is configured to issue various commands for performingsequencing-by-synthesis processes. In SBS, extension of a nucleic acidprimer along a nucleic acid template is monitored to determine thesequence of nucleotides in the template. The underlying chemical processcan be polymerization (e.g. as catalyzed by a polymerase enzyme) orligation (e.g. catalyzed by a ligase enzyme). In a particularpolymerase-based SBS embodiment, fluorescently labeled nucleotides areadded to a primer (thereby extending the primer) in a template dependentfashion such that detection of the order and type of nucleotides addedto the primer can be used to determine the sequence of the template. Forexample, to initiate a first SBS cycle, commands can be given to deliverone or more labeled nucleotides, DNA polymerase, etc., into/through aflow cell that houses an array of nucleic acid templates. The nucleicacid templates may be located at corresponding reaction sites. Thosereaction sites where primer extension causes a labeled nucleotide to beincorporated can be detected through an imaging event. During an imagingevent, the illumination system 111 may provide an excitation light tothe reaction sites. Optionally, the nucleotides can further include areversible termination property that terminates further primer extensiononce a nucleotide has been added to a primer. For example, a nucleotideanalog having a reversible terminator moiety can be added to a primersuch that subsequent extension cannot occur until a deblocking agent isdelivered to remove the moiety. Thus, for embodiments that usereversible termination a command can be given to deliver a deblockingreagent to the flow cell (before or after detection occurs). One or morecommands can be given to effect wash(es) between the various deliverysteps. The cycle can then be repeated n times to extend the primer by nnucleotides, thereby detecting a sequence of length n. Exemplarysequencing techniques are described, for example, in Bentley et al.,Nature 456:53-59 (2008), WO 04/018497; U.S. Pat. No. 7,057,026; WO91/06678; WO 07/123744; U.S. Pat. Nos. 7,329,492; 7,211,414; 7,315,019;7,405,281, and US 2008/0108082, each of which is incorporated herein byreference.

For the nucleotide delivery step of an SBS cycle, either a single typeof nucleotide can be delivered at a time, or multiple differentnucleotide types (e.g. A, C, T and G together) can be delivered. For anucleotide delivery configuration where only a single type of nucleotideis present at a time, the different nucleotides need not have distinctlabels since they can be distinguished based on temporal separationinherent in the individualized delivery. Accordingly, a sequencingmethod or apparatus can use single color detection. For example, anexcitation source need only provide excitation at a single wavelength orin a single range of wavelengths. For a nucleotide deliveryconfiguration where delivery results in multiple different nucleotidesbeing present in the flow cell at one time, sites that incorporatedifferent nucleotide types can be distinguished based on differentfluorescent labels that are attached to respective nucleotide types inthe mixture. For example, four different nucleotides can be used, eachhaving one of four different fluorophores. In one embodiment, the fourdifferent fluorophores can be distinguished using excitation in fourdifferent regions of the spectrum. For example, four differentexcitation radiation sources can be used. Alternatively, fewer than fourdifferent excitation sources can be used, but optical filtration of theexcitation radiation from a single source can be used to producedifferent ranges of excitation radiation at the flow cell.

In some embodiments, fewer than four different colors can be detected ina mixture having four different nucleotides. For example, pairs ofnucleotides can be detected at the same wavelength, but distinguishedbased on a difference in intensity for one member of the pair comparedto the other, or based on a change to one member of the pair (e.g. viachemical modification, photochemical modification or physicalmodification) that causes apparent signal to appear or disappearcompared to the signal detected for the other member of the pair.Exemplary apparatus and methods for distinguishing four differentnucleotides using detection of fewer than four colors are described forexample in U.S. Pat. App. Ser. Nos. 61/538,294 and 61/619,878, which areincorporated herein by reference their entireties. U.S. application Ser.No. 13/624,200, which was filed on Sep. 21, 2012, is also incorporatedby reference in its entirety.

The plurality of protocol modules may also include a sample-preparation(or generation) module 137 that is configured to issue commands to thefluidic control system 106 and the temperature control system 110 foramplifying a product within the biosensor 102. For example, thebiosensor 102 may be engaged to the bioassay system 100. Theamplification module 137 may issue instructions to the fluidic controlsystem 106 to deliver necessary amplification components to reactionchambers within the biosensor 102. In other embodiments, the reactionsites may already contain some components for amplification, such as thetemplate DNA and/or primers. After delivering the amplificationcomponents to the reaction chambers, the amplification module 137 mayinstruct the temperature control system 110 to cycle through differenttemperature stages according to known amplification protocols. In someembodiments, the amplification and/or nucleotide incorporation isperformed isothermally.

The SBS module 136 may issue commands to perform bridge PCR whereclusters of clonal amplicons are formed on localized areas within achannel of a flow cell. After generating the amplicons through bridgePCR, the amplicons may be “linearized” to make single stranded templateDNA, or sstDNA, and a sequencing primer may be hybridized to a universalsequence that flanks a region of interest. For example, a reversibleterminator-based sequencing by synthesis method can be used as set forthabove or as follows.

Each sequencing cycle can extend a sstDNA by a single base which can beaccomplished for example by using a modified DNA polymerase and amixture of four types of nucleotides. The different types of nucleotidescan have unique fluorescent labels, and each nucleotide can further havea reversible terminator that allows only a single-base incorporation tooccur in each cycle. After a single base is added to the sstDNA,excitation light may be incident upon the reaction sites and fluorescentemissions may be detected. After detection, the fluorescent label andthe terminator may be chemically cleaved from the sstDNA. Anothersimilar sequencing cycle may follow. In such a sequencing protocol, theSBS module 136 may instruct the fluidic control system 106 to direct aflow of reagent and enzyme solutions through the biosensor 102.Exemplary reversible terminator-based SBS methods which can be utilizedwith the apparatus and methods set forth herein are described in USPatent Application Publication No. 2007/0166705 A1, US PatentApplication Publication No. 2006/0188901 A1, U.S. Pat. No. 7,057,026, USPatent Application Publication No. 2006/0240439 A1, US PatentApplication Publication No. 2006/0281109 A1, PCT Publication No. WO05/065814, US Patent Application Publication No. 2005/0100900 A1, PCTPublication No. WO 06/064199 and PCT Publication No. WO 07/010251, eachof which is incorporated herein by reference in its entirety. Exemplaryreagents for reversible terminator-based SBS are described in U.S. Pat.Nos. 7,541,444; 7,057,026; 7,414,116; 7,427,673; 7,566,537; 7,592,435and WO 07/135368, each of which is incorporated herein by reference inits entirety.

In some embodiments, the amplification and SBS modules may operate in asingle assay protocol where, for example, template nucleic acid isamplified and subsequently sequenced within the same cartridge.

The bioassay system 100 may also allow the user to reconfigure an assayprotocol. For example, the bioassay system 100 may offer options to theuser through the user interface 114 for modifying the determinedprotocol. For example, if it is determined that the biosensor 102 is tobe used for amplification, the bioassay system 100 may request atemperature for the annealing cycle. Furthermore, the bioassay system100 may issue warnings to a user if a user has provided user inputs thatare generally not acceptable for the selected assay protocol.

FIG. 3 is a block diagram of an exemplary workstation 200 for biologicalor chemical analysis in accordance with one embodiment. The workstation200 may have similar features, systems, and assemblies as the bioassaysystem 100 described above. For example, the workstation 200 may have afluidic control system, such as the fluidic control system 106 (FIG. 1),that is fluidicly coupled to a biosensor (or cartridge) 235 through afluid network 238. The fluid network 238 may include a reagent cartridge240, a valve block 242, a main pump 244, a debubbler 246, a 3-way valve248, a flow restrictor 250, a waste removal system 252, and a purge pump254. In particular embodiments, most of the components or all of thecomponents described above are within a common workstation housing (notshown). Although not shown, the workstation 200 may also include anillumination system, such as the illumination system 111, that isconfigured to provide an excitation light to the reaction sites.

A flow of fluid is indicated by arrows along the fluid network 238. Forexample, reagent solutions may be removed from the reagent cartridge 240and flow through the valve block 242. The valve block 242 may facilitatecreating a zero-dead volume of the fluid flowing to the cartridge 235from the reagent cartridge 240. The valve block 242 can select or permitone or more liquids within the reagent cartridge 240 to flow through thefluid network 238. For example, the valve block 242 can include solenoidvalves that have a compact arrangement. Each solenoid valve can controlthe flow of a fluid from a single reservoir bag. In some embodiments,the valve block 242 can permit two or more different liquids to flowinto the fluid network 238 at the same time thereby mixing the two ormore different liquids. After leaving the valve block 242, the fluid mayflow through the main pump 244 and to the debubbler 246. The debubbler246 is configured to remove unwanted gases that have entered or beengenerated within the fluid network 238.

From the debubbler 246, fluid may flow to the 3-way valve 248 where thefluid is either directed to the cartridge 235 or bypassed to the wasteremoval system 252. A flow of the fluid within the cartridge 235 may beat least partially controlled by the flow restrictor 250 locateddownstream from the cartridge 235. Furthermore, the flow restrictor 250and the main pump 244 may coordinate with each other to control the flowof fluid across reaction sites and/or control the pressure within thefluid network 238. Fluid may flow through the cartridge 235 and onto thewaste removal system 252. Optionally, fluid may flow through the purgepump 254 and into, for example, a waste reservoir bag within the reagentcartridge 240.

Also shown in FIG. 3, the workstation 200 may include a temperaturecontrol system, such as the temperature control system 110, that isconfigured to regulate or control a thermal environment of the differentcomponents and sub-systems of the workstation 200. The temperaturecontrol system 110 can include a reagent cooler 264 that is configuredto control the temperature requirements of various fluids used by theworkstation 200, and a thermocycler 266 that is configured to controlthe temperature of a cartridge 235. The thermocycler 266 can include athermal element (not shown) that interfaces with the cartridge.

Furthermore, the workstation 200 may include a system controller or SBSboard 260 that may have similar features as the system controller 104described above. The SBS board 260 may communicate with the variouscomponents and sub-systems of the workstation 200 as well as thecartridge 235. Furthermore, the SBS board 260 may communicate withremote systems to, for example, store data or receive commands from theremote systems. The workstation 200 may also include a touch screen userinterface 262 that is operatively coupled to the SBS board 260 through asingle-board computer (SBC) 272. The workstation 200 may also includeone or more user accessible data communication ports and/or drives. Forexample a workstation 200 may include one or more universal serial bus(USB) connections for computer peripherals, such as a flash or jumpdrive, a compact-flash (CF) drive and/or a hard drive 270 for storinguser data in addition to other software.

FIG. 4 is a perspective view of a workstation 300 and a cartridge 302that may include one or more biosensors (not shown) as described herein.The workstation 300 may include similar components as described abovewith respect to the bioassay system 100 and the workstation 200 and mayoperate in a similar manner. For example, the workstation 300 mayinclude a workstation housing 304 and a system receptacle 306 that isconfigured to receive and engage the cartridge 302. The systemreceptacle may at least one of fluidically or electrically engage thecartridge 302. The workstation housing 304 may hold, for example, asystem controller, a fluid storage system, a fluidic control system, anda temperature control system as described above. In FIG. 4, theworkstation 300 does not include a user interface or display that iscoupled to the workstation housing 304. However, a user interface may becommunicatively coupled to the housing 304 (and the components/systemstherein) through a communication link. Thus, the user interface and theworkstation 300 may be remotely located with respect to each other.Together, the user interface and the workstation 300 (or a plurality ofworkstations) may constitute a bioassay system.

As shown, the cartridge 302 includes a cartridge housing 308 having atleast one port 310 that provides access to an interior of the cartridgehousing 308. For example, a solution that is configured to be used inthe cartridge 302 during the controlled reactions may be insertedthrough the port 310 by a technician or by the workstation 300. Thesystem receptacle 306 and the cartridge 302 may be sized and shapedrelative to each other such that the cartridge 302 may be inserted intoa receptacle cavity (not shown) of the system receptacle 306.

FIG. 5 is a front view of a rack assembly 312 having a cabinet orcarriage 314 with a plurality of the workstations 300 loaded thereon.The cabinet 314 may include one or more shelves 316 that define one ormore reception spaces 318 configured to receive one or more workstations300. Although not shown, the workstations 300 may be communicativelycoupled to a communication network that permits a user to controloperation of the workstations 300. In some embodiments, a bioassaysystem includes a plurality of workstations, such as the workstations300, and a single user interface configured to control operation of themultiple workstations.

FIG. 6 illustrates various features of the cartridge 302 (FIG. 4) inaccordance with one embodiment. As shown, the cartridge 302 may includea sample assembly 320, and the system receptacle 306 may include a lightassembly 322. Stage 346 shown in FIG. 6 represents the spatialrelationship between the first and second sub-assemblies 320 and 322when they are separate from each other. At stage 348, the first andsecond sub-assemblies 320 and 322 are joined together. The cartridgehousing 308 (FIG. 4) may enclose the joined first and secondsub-assemblies 320 and 322.

In the illustrated embodiment, the first sub-assembly 320 includes abase 326 and a reaction-component body 324 that is mounted onto the base326. Although not shown, one or more biosensors may be mounted to thebase 326 in a recess 328 that is defined, at least in part, by thereaction-component body 324 and the base 326. For example, at least fourbiosensors may be mounted to the base 326. In some embodiments, the base326 is a printed circuit board having circuitry that enablescommunication between the different components of the cartridge and theworkstation 300 (FIG. 4). For example, the reaction-component body 324may include a rotary valve 330 and reagent reservoirs 332 that arefluidically coupled to the rotary valve 330. The reaction-component body324 may also include additional reservoirs 334.

The second sub-assembly 322 includes a light assembly 336 that includesa plurality of light directing channels 338. Each light directingchannel 338 is optically coupled to a light source (not shown), such asa light-emitting diode (LED). The light source(s) are configured toprovide an excitation light that is directed by the light directingchannels 338 onto the biosensors. In alternative embodiments, thecartridge may not include a light source(s). In such embodiments, thelight source(s) may be located in the workstation 300. When thecartridge is inserted into the system receptacle 306 (FIG. 4), thecartridge 302 may align with the light source(s) so that the biosensorsmay be illuminated.

Also shown in FIG. 6, the second sub-assembly 322 includes a cartridgepump 340 that is fluidically coupled to ports 342 and 344. When thefirst and second sub-assemblies 320 and 322 are joined together, theport 342 is coupled to the rotary valve 330 and the port 344 is coupledto the other reservoirs 334. The cartridge pump 340 may be activated todirect reaction components from the reservoirs 332 and/or 334 to thebiosensors according to a designated protocol.

FIG. 7 illustrates a cross-section of a portion of an exemplarybiosensor 400 formed in accordance with one embodiment. The biosensor400 may include similar features as the biosensor 102 (FIG. 1) describedabove and may be used in, for example, the cartridge 302 (FIG. 4). Asshown, the biosensor 400 may include a flow cell 402 that is coupleddirectly or indirectly to a detection device 404. The flow cell 402 maybe mounted to the detection device 404. In the illustrated embodiment,the flow cell 402 is affixed directly to the detection device 404through one or more securing mechanisms (e.g., adhesive, bond,fasteners, and the like). In some embodiments, the flow cell 402 may beremovably coupled to the detection device 404.

In the illustrated embodiment, the detection device 404 includes adevice base 425. In particular embodiments, the device base 425 includesa plurality of stacked layers (e.g., silicon layer, dielectric layer,metal-dielectric layers, etc.). The device base 425 may include a sensorarray 424 of light sensors 440, a guide array 426 of light guides 462,and a reaction array 428 of reaction recesses 408 that havecorresponding reaction sites 414. In certain embodiments, the componentsare arranged such that each light sensor 440 aligns with a single lightguide 462 and a single reaction site 414. However, in other embodiments,a single light sensor 440 may receive photons through more than onelight guide 462 and/or from more than one reaction site 414. As usedherein, a single light sensor may include one pixel or more than onepixel.

Moreover, it is noted that the term “array” or “sub-array” does notnecessarily include each and every item of a certain type that thedetection device may have. For example, the sensor array 424 may notinclude each and every light sensor in the detection device 404.Instead, the detection device 404 may include other light sensors (e.g.,other array(s) of light sensors). As another example, the guide array426 may not include each and every light guide of the detection device.Instead, there may be other light guides that are configured differentlythan the light guides 462 or that have different relationships withother elements of the detection device 404. As such, unless explicitlyrecited otherwise, the term “array” may or may not include all suchitems of the detection device.

In the illustrated embodiment, the flow cell 402 includes a sidewall 406and a flow cover 410 that is supported by the sidewall 406 and othersidewalls (not shown). The sidewalls are coupled to the detector surface412 and extend between the flow cover 410 and the detector surface 412.In some embodiments, the sidewalls are formed from a curable adhesivelayer that bonds the flow cover 410 to the detection device 404.

The flow cell 402 is sized and shaped so that a flow channel 418 existsbetween the flow cover 410 and the detection device 404. As shown, theflow channel 418 may include a height H₁. By way of example only, theheight H₁ may be between about 50-400 μm (microns) or, moreparticularly, about 80-200 μm. In the illustrated embodiment, the heightH₁ is about 100 μm. The flow cover 410 may include a material that istransparent to excitation light 401 propagating from an exterior of thebiosensor 400 into the flow channel 418. As shown in FIG. 7, theexcitation light 401 approaches the flow cover 410 at a non-orthogonalangle. However, this is only for illustrative purposes as the excitationlight 401 may approach the flow cover 410 from different angles.

Also shown, the flow cover 410 may include inlet and outlet ports 420,422 that are configured to fluidically engage other ports (not shown).For example, the other ports may be from the cartridge 302 (FIG. 4) orthe workstation 300 (FIG. 4). The flow channel 418 is sized and shapedto direct a fluid along the detector surface 412. The height H₁ andother dimensions of the flow channel 418 may be configured to maintain asubstantially even flow of a fluid along the detector surface 412. Thedimensions of the flow channel 418 may also be configured to controlbubble formation.

The sidewalls 406 and the flow cover 410 may be separate components thatare coupled to each other. In other embodiments, the sidewalls 406 andthe flow cover 410 may be integrally formed such that the sidewalls 406and the flow cover 410 are formed from a continuous piece of material.By way of example, the flow cover 410 (or the flow cell 402) maycomprise a transparent material, such as glass or plastic. The flowcover 410 may constitute a substantially rectangular block having aplanar exterior surface and a planar inner surface that defines the flowchannel 418. The block may be mounted onto the sidewalls 406.Alternatively, the flow cell 402 may be etched to define the flow cover410 and the sidewalls 406. For example, a recess may be etched into thetransparent material. When the etched material is mounted to thedetection device 404, the recess may become the flow channel 418.

The detection device 404 has a detector surface 412 that may befunctionalized (e.g., chemically or physically modified in a suitablemanner for conducting designated reactions). For example, the detectorsurface 412 may be functionalized and may include a plurality ofreaction sites 414 having one or more biomolecules immobilized thereto.The detector surface 412 has an array of reaction recesses or open-sidedreaction chambers 408. Each of the reaction recesses 408 may include oneor more of the reaction sites 414. The reaction recesses 408 may bedefined by, for example, an indent or change in depth along the detectorsurface 412. In other embodiments, the detector surface 412 may besubstantially planar.

As shown in FIG. 7, the reaction sites 414 may be distributed in apattern along the detector surface 412. For instance, the reactionssites 414 may be located in rows and columns along the detector surface412 in a manner that is similar to a microarray. However, it isunderstood that various patterns of reaction sites may be used. Thereaction sites may include biological or chemical substances that emitlight signals. For example, the biological or chemical substances of thereactions sites may generate light emissions in response to theexcitation light 401. In particular embodiments, the reaction sites 414include clusters or colonies of biomolecules (e.g., oligonucleotides)that are immobilized on the detector surface 412.

FIG. 8 is an enlarged cross-section of the detection device 404 showingvarious features in greater detail. More specifically, FIG. 8 shows asingle light sensor 440, a single light guide 462 for directing lightemissions toward the light sensor 440, and associated circuitry 446 fortransmitting signals based on the light emissions (e.g., photons)detected by the light sensor 440. It is understood that the other lightsensors 440 of the sensor array 424 (FIG. 7) and associated componentsmay be configured in an identical or similar manner. It is alsounderstood, however, the detection device 404 is not required to bemanufactured identically or uniformly throughout. Instead, one or morelight sensors 440 and/or associated components may be manufactureddifferently or have different relationships with respect to one another.

The circuitry 446 may include interconnected conductive elements (e.g.,conductors, traces, vias, interconnects, etc.) that are capable ofconducting electrical current, such as the transmission of data signalsthat are based on detected photons. For example, in some embodiments,the circuitry 446 may be similar to or include a microcircuitarrangement, such as the microcircuit arrangement described in U.S. Pat.No. 7,595,883, which is incorporated herein by reference in theentirety. The detection device 404 and/or the device base 425 maycomprise an integrated circuit having a planar array of the lightsensors 440. The circuitry 446 formed within the detection device 425may be configured for at least one of signal amplification,digitization, storage, and processing. The circuitry may collect andanalyze the detected light emissions and generate data signals forcommunicating detection data to a bioassay system. The circuitry 446 mayalso perform additional analog and/or digital signal processing in thedetection device 404.

The device base 425 may be manufactured using integrated circuitmanufacturing processes, such as processes used to manufacturecomplementary-metal-oxide semiconductors (CMOSs). For example, thedevice base 425 may include a plurality of stacked layers 431-437including a sensor layer or base 431, which is a silicon layer or waferin the illustrated embodiment. The sensor layer 431 may include thelight sensor 440 and gates 441-443 that are formed with the sensor layer431. The gates 441-443 are electrically coupled to the light sensor 440.When the detection device 404 is fully formed as shown in FIGS. 7 and 8,the light sensor 440 may be electrically coupled to the circuitry 446through the gates 441-443.

As used herein, the term “layer” is not limited to a single continuousbody of material unless otherwise noted. For example, the sensor layer431 may include multiple sub-layers that are different materials and/ormay include coatings, adhesives, and the like. Furthermore, one or moreof the layers (or sub-layers) may be modified (e.g., etched, depositedwith material, etc.) to provide the features described herein.

In some embodiments, each light sensor 440 has a detection area that isless than about 50 μm². In particular embodiments, the detection area isless than about 10 μm². In more particular embodiments, the detectionarea is about 2 μm². In such cases, the light sensor 440 may constitutea single pixel. An average read noise of each pixel in a light sensor440 may be, for example, less than about 150 electrons. In moreparticular embodiments, the read noise may be less than about 5electrons. The resolution of the array of light sensors 440 may begreater than about 0.5 megapixels (Mpixels). In more specificembodiments, the resolution may be greater than about 5 Mpixels and,more particularly, greater than about 10 Mpixels.

The device layers also include a plurality of metal-dielectric layers432-437, which are hereinafter referred to as substrate layers. In theillustrated embodiment, each of the substrate layers 432-437 includesmetallic elements (e.g., W (tungsten), Cu (copper), or Al (aluminum))and dielectric material (e.g., SiO₂). Various metallic elements anddielectric material may be used, such as those suitable for integratedcircuit manufacturing. However, in other embodiments, one or more of thesubstrate layers 432-437 may include only dielectric material, such asone or more layers of SiO₂.

With respect to the specific embodiment shown in FIG. 8, the firstsubstrate layer 432 may include metallic elements referred to as M1 thatare embedded within dielectric material (e.g., SiO₂). The metallicelements M1 comprise, for example, W (tungsten). The metallic elementsM1 extend entirely through the substrate layer 432 in the illustratedembodiment. The second substrate layer 433 includes metallic elements M2and dielectric material as well as a metallic interconnects (M2/M3). Thethird substrate layer 434 includes metallic elements M3 and metalinterconnects (M3/M4). The fourth substrate layer 435 also includesmetallic elements M4. The device base 425 also includes fifth and sixthsubstrate layers 436, 437, which are described in greater detail below.

As shown, the metallic elements and interconnects are connected to eachother to form at least a portion of the circuitry 446. In theillustrated embodiment, the metallic elements M1, M2, M3, M4 include W(tungsten), Cu (copper), and/or aluminum (Al) and the metalinterconnects M2/M3 and M3/M4 include W (tungsten), but it is understoodthat other materials and configurations may be used. It is also notedthat the device base 425 and the detection device 404 shown in FIGS. 7and 8 are for illustrative purposes only. For example, other embodimentsmay include fewer or additional layers than those shown in FIGS. 7 and 8and/or different configurations of metallic elements.

In some embodiments, the detection device 404 includes a shield layer450 that extends along an outer surface 464 of the device base 425. Inthe illustrated embodiment, the shield layer 450 is deposited directlyalong the outer surface 464 of the substrate layer 437. However, anintervening layer may be disposed between the substrate layer 437 andthe shield layer 450 in other embodiments. The shield layer 450 mayinclude a material that is configured to block, reflect, and/orsignificantly attenuate the light signals that are propagating from theflow channel 418. The light signals may be the excitation light 401and/or the light emissions 466 (shown in FIG. 9). By way of exampleonly, the shield layer 450 may comprise tungsten (W).

As shown in FIG. 8, the shield layer 450 includes an aperture or opening452 therethrough. The shield layer 450 may include an array of suchapertures 452. In some embodiments, the shield layer 450 may extendcontinuously between adjacent apertures 452. As such, the light signalsfrom the flow channel 418 may be blocked, reflected, and/orsignificantly attenuated to prevent detection of such light signals bythe light sensors 440. However, in other embodiments, the shield layer450 does not extend continuously between the adjacent apertures 452 suchthen one or more openings other than the apertures 452 exits in theshield layer 450.

The detection device 404 may also include a passivation layer 454 thatextends along the shield layer 450 and across the apertures 452. Theshield layer 450 may extend over the apertures 452 thereby directly orindirectly covering the apertures 452. The shield layer 450 may belocated between the passivation layer 454 and the device base 425. Anadhesive or promoter layer 458 may be located therebetween to facilitatecoupling the passivation and shield layers 454, 450. The passivationlayer 454 may be configured to protect the device base 425 and theshield layer 450 from the fluidic environment of the flow channel 418.

In some cases, the passivation layer 454 may also be configured toprovide a solid surface (i.e., the detector surface 412) that permitsbiomolecules or other analytes-of-interest to be immobilized thereon.For example, each of the reaction sites 414 may include a cluster ofbiomolecules that are immobilized to the detector surface 412 of thepassivation layer 454. Thus, the passivation layer 454 may be formedfrom a material that permits the reaction sites 414 to be immobilizedthereto. The passivation layer 454 may also comprise a material that isat least transparent to a desired fluorescent light. By way of example,the passivation layer 454 may include silicon nitride (Si₃N₄) and/orsilica (SiO₂). However, other suitable material(s) may be used. Inaddition, the passivation layer 454 may be physically or chemicallymodified to facilitate immobilizing the biomolecules and/or tofacilitate detection of the light emissions.

In the illustrated embodiment, a portion of the passivation layer 454extends along the shield layer 450 and a portion of the passivationlayer 454 extends directly along filter material 460 of a light guide462. The reaction recess 408 may be formed directly over the light guide462. In some cases, prior to the passivation layer 454 being depositedalong the shield layer 450 or adhesion layer 458, a base hole or cavity456 may be formed within the device base 425. For example, the devicebase 425 may be etched to form an array of the base holes 456. Inparticular embodiments, the base hole 456 is an elongated space thatextends from proximate the aperture 452 toward the light sensor 440. Thebase hole may extend lengthwise along a central longitudinal axis 468. Athree-dimensional shape of the base hole 456 may be substantiallycylindrical or frustro-conical in some embodiments such that across-section taken along a plane that extends into the page of FIG. 8is substantially circular. The longitudinal axis 468 may extend througha geometric center of the cross-section. However, other geometries maybe used in alternative embodiments. For example, the cross-section maybe substantially square-shaped or octagonal.

The filter material 460 may be deposited within the base hole 456 afterthe base hole 456 is formed. The filter material 460 may form (e.g.,after curing) a light guide 462. The light guide 462 is configured tofilter the excitation light 401 and permit the light emissions 466 topropagate therethrough toward the corresponding light sensor 440. Thelight guide 462 may be, for example, an organic absorption filter. Byway of specific example only, the excitation light may be about 532 nmand the light emissions may be about 570 nm or more.

In some cases, the organic filter material may be incompatible withother materials of the biosensor. For example, organic filter materialmay have a coefficient of thermal expansion that causes the filtermaterial to significantly expand. Alternatively or in addition to, thefilter material may be unable to sufficiently adhere to certain layers,such as the shield layer (or other metal layers). Expansion of thefilter material may cause mechanical stress on the layers that areadjacent to the filter material or structurally connected to the filtermaterial. In some cases, the expansion may cause cracks or otherunwanted features in the structure of the biosensor. As such,embodiments set forth herein may limit the degree to which the filtermaterial expands and/or the degree to which the filter material is incontact with other layers. For example, the filter material of differentlight guides may be isolated from each other by the passivation layer.In such embodiments, the filter material may not contact the metallayer(s). Moreover, the passivation layer may resist expansion and/orpermit some expansion while reducing generation of unwanted structuralfeatures (e.g., cracks).

The light guide 462 may be configured relative to surrounding materialof the device base 425 (e.g., the dielectric material) to form alight-guiding structure. For example, the light guide 462 may have arefractive index of about 2.0 so that the light emissions aresubstantially reflected at an interface between the light guide 462 andthe material of the device base 425. In certain embodiments, the lightguide 462 is configured such that the optical density (OD) or absorbanceof the excitation light is at least about 4 OD. More specifically, thefilter material may be selected and the light guide 462 may bedimensioned to achieve at least 4 OD. In more particular embodiments,the light guide 462 may be configured to achieve at least about 5 OD orat least about 6 OD. Other features of the biosensor 400 may beconfigured to reduce electrical and optical crosstalk.

FIG. 9 illustrates an enlarged view of the detector surface 412 andportions of the detection device 404 (FIG. 7) that are located proximateto the detector surface 412. More specifically, the passivation layer454, the adhesion layer 458, the shield layer 450, and the light guide462 are shown in FIG. 9. Each of the layers may have a outer (top)surface or an inner (bottom) surface and may extend along an adjacentlayer at an interface. In some embodiments, the detector surface 412 isconfigured to form the reaction recess 408 proximate to the aperture452. The reaction recess 408 may be, for example, an indent, pit, well,groove, or open-sided chamber or channel. Alternatively, the detectorsurface 412 may be planar without the recesses shown in FIGS. 7-9. Asshown, the aperture 452 is defined by an aperture or layer edge 504. Thelayer edge 504 faces radially inward toward the longitudinal axis 468.

The detector surface 412 may include an elevated portion 502 and thereaction recess 408 may include a base surface 490. The base surface 490may extend substantially parallel to the shield layer 450. The detectorsurface 412 may also include a side surface 492 that extendssubstantially orthogonal to the base surface 490 and the elevatedportion 502 of the detector surface 412. The side surface 492 may definea periphery of the reaction recess 408. Although the elevated portion502, the base surface 490, and the side surface 492 are referenced asseparate surfaces it is understood that the surfaces may be portions ofthe detector surface 412. Moreover, it is understood that, due tomanufacturing tolerances, the surfaces may not have be readily distinct.For example, in other embodiments, the base surface 490 and the sidesurface 492 may be substantially a single surface with a concave shape.

The base surface 490 may represent (or include a point that represents)a deepest portion of the passivation layer 454 along the detectorsurface 412 within the reaction recess 408. For example, the elevatedportion 502 may extend along a surface plane P₁ and the base surface 490may extend along a surface plane P₂. As shown, the surface planes P₁ andP₂ are offset with respect to each other by a depth or distance D₁. Thesurface plane P₂ is closer to the light guide 462 or the light sensor440 (FIG. 7) than the surface plane P₁. In the illustrated embodiment,the depth D₁ of the base surface 490 is substantially continuous due tothe base surface 490 being substantially planar. In other embodiment,however, the depth Di may vary. For example, the base surface 490 mayhave a concave shape with the depth increasing as the base surface 490extends toward a center or middle thereof.

The reaction recess 408 may extend toward or be located within theaperture 452. For instance, at least a portion of the base surface 490may reside within the aperture 452. The shield layer 450 may have anouter surface 506 that faces the passivation layer 454 and an innersurface 508 that faces the device base 425. The outer surface 506 mayextend along a surface plane P₃, and the inner surface 508 may extendalong a surface plane P₄. The distance between the surface planes P₃ andP₄ may represent a thickness of the shield layer 450. As shown, thesurface plane P₃ may be located between the surface planes P₁, P₂. Assuch, the base surface 490 extends within the aperture 452 as defined bythe layer edge 504. In other embodiments, however, the surface plane P₂may be located above the surface plane P₃ such that the base surface 490does not reside within the aperture 452. Moreover, in some embodiments,the surface plane P₂ may be located below the surface plane P₄ such thatbase surface 490 is located below the aperture 452.

The passivation layer 454 includes the detector surface 412 and an innersurface 510 that extends along the outer surface 506 of the shield layer450 at an interface 512. In some embodiments, the adhesion layer 458 mayextend along and define the interface 512 between the shield layer 450and the passivation layer 454.

In the illustrated embodiment, the passivation layer 454 extendsdirectly along the light guide 462. More specifically, the inner surface510 of the passivation layer 454 may directly engage a material surface514 of the light guide 462. As used herein, the phrase “directly engage”and the like may include the two layers directly contacting each otheror the two layers being bonded to each other through the use of anadhesion promoter material(s). The light guide 462 has an input region472 that includes the material surface 514. The input region 472 mayrepresent a portion of the light guide 462 that initially receives thelight emissions.

The inner surface 510 may directly engage the material surface 514 at aninterface 516. The interface 516 may represent a material level of thefilter material 460 that is deposited within the guide cavity 456 (FIG.7). In the illustrated embodiment, the interface 516 is substantiallyplanar such that the interface 516 extends along an interface plane P₅.The interface plane P₅ may extend substantially parallel to one or moreof the surface planes P₁, P₂, P₃, P₄. In other embodiments, however, theinterface 516 may have a concave shape such that the interface 516 bowstoward the light sensor 440 (FIG. 8) or in an opposite direction awayfrom the light sensor 440.

The passivation layer 454 may fill a void generated when the aperture452 is formed. Thus, in some embodiments, the passivation layer 454 maybe located within or reside in the aperture 452. In particularembodiments, the interface 516 may be located a depth D₂ into the devicebase 425. In particular embodiments, the depth D₂ may be configured suchthat the interface 516 is located below the aperture 452 as shown inFIG. 8. In such embodiments, the passivation layer 454 may isolate(e.g., separate) the filter material 460 and the shield layer 450. Suchembodiments may be suitable when the filter material 460 and the shieldlayer 450 are incompatible such that cracks or other unwanted featuresmay develop during manufacture of usage of the biosensor 400 (FIG. 7).In other embodiments, at least a portion of the interface 516 may belocated within the aperture 452.

Also shown in FIG. 9, the passivation layer 454 may form a joint orcorner region 519. The joint region 519 may include the side surface 492and extend around the longitudinal axis 468. The joint region 519 mayinclude a relatively thicker portion of the passivation layer 454 thatextends from the elevated portion 502 to the inner surface 510 at thematerial interface 516 (or between the surface plane P₁ and theinterface plane P₅). The dimensions of the joint region 519 may resistmechanical stresses caused by expansion of the filter material 460during manufacture of the biosensor 400 and/or during thermal cyclingthat may occur during designated protocols (e.g., SBS sequencing). Asshown, the thickness between the surface plane P₁ and the interfaceplane P₅ is more than twice the thickness between the elevated portion502 of the detector surface 412 and the interface 512.

The reaction site 414 may include biological or chemical substances,which are generally represented as dots 520 in FIG. 9. The biological orchemical substances may be immobilized to the detector surface 412 or,more specifically, the base and side surfaces 490, 492. In particularembodiments, the reaction site 414 is located proximate to the aperture452 so that light emissions propagate through the passivation layer 454,through the aperture 452, and into the input region 472 of the lightguide 462.

In some embodiments, the reaction sites 414 or the biological orchemical substances 520 therein may be patterned such that the reactionsites 414 or substances 520 have predetermined locations. For example,after the passivation layer 454 is applied, the reaction sites 414 orportions thereof may be patterned onto the passivation layer 454. In theillustrate embodiment, each aperture 452 is associated with a singlereaction site 414 such that the light emissions from the reaction site414 are directed toward the corresponding light sensor 440. Thebiological or chemical substances 520 in a single reaction site 414 maybe similar or identical (e.g., a colony of oligonucleotides that have acommon sequence). However, in other embodiments, more than one reactionsite 414 may correspond to one of the apertures 452.

In particular embodiments, the reaction sites 414 may include pads ormetal regions that are described in U.S. Provisional Application No.61/495,266, filed on Jun. 9, 2011, and U.S. Provisional Application No.61/552,712, filed on Oct. 28, 2011. Each of the U.S. ProvisionalApplication No. 61/495,266 (the '266 Application) and the U.S.Provisional Application No. 61/552,712 (the '712 Application) isincorporated herein by reference in its entirety. In some embodiments,the reaction sites 414 may be fabricated after the flow cell 402 (FIG.7) is manufactured on the detection device 404.

In the illustrated embodiment, the reaction site 414 includes a colonyof oligonucleotides 520 in which the oligonucleotides have aneffectively common sequence. In such embodiments, each of theoligonucleotides may generate common light emissions when the excitationlight 401 is absorbed by the fluorophors incorporated within theoligonucleotides. As shown, the light emissions 466 may emit in alldirections (e.g., isotropically) such that, for example, a portion ofthe light is directed into the light guide 462, a portion of the lightis directed to reflect off the shield layer 450, and a portion of thelight is directed into the flow channel 418 or the passivation layer454. For the portion that is directed into the light guide 462,embodiments described herein may be configured to facilitate detectionof the photons.

Also shown in FIG. 9, the device base 425 may include peripheralcrosstalk shields 522, 524 located within the device base 425. Thecrosstalk shields 522, 524 may be positioned relative to the light guide462 and configured so that the crosstalk shields 522, 524 block orreflect light signals propagating out of the light guide 462. The lightsignals may include the excitation light 401 that has been reflected orrefracted and/or the light emissions 466 generated at or proximate tothe detector surface 412. In some embodiments, the crosstalk shields522, 524 may also directly block the excitation light 401 from the flowchannel 418. As such, the crosstalk shields 522, 524 may reducedetection of unwanted light signals. For example, the crosstalk shields522, 524 may reduce optical crosstalk between adjacent light sensors 440and/or may improve collection efficiency of the corresponding lightsensor 440. The crosstalk shields 522, 524 may be, for example, metallicelements that are fabricated during the manufacture of the device base425. In some embodiments, the processes used to fabricate the M1, M2,M3, M2/M3, and M3/M4 elements of the circuitry 446 (FIG. 8) may be thesame as or similar to the processes that fabricate the crosstalk shields522, 524. For example, the crosstalk shields 522, 524 may be locatedwithin dielectric material (e.g., dielectric layers) of the device base425 and comprise the same material that is used to fabricate thecircuitry 446 (e.g., one or more of the materials used to fabricate theM1, M2, M3, M2/M3, and M3/M4 elements). Although not shown, in somecases, the different stages of CMOS manufacture may include forming themetallic elements that will transmit data signals while also forming thecrosstalk shields.

Although the crosstalk shields 522, 524 may be manufactured in a similarmanner as the circuitry 446, the crosstalk shields 522, 524 may beelectrically separate from the circuitry 446. In other words, for someembodiments, the crosstalk shields 522, 524 may not transmit datasignals. In other embodiments, however, the crosstalk shields 522, 524may be traces or other metallic elements that are configured to transmitdata signals. As also shown in FIG. 9, the crosstalk shields 522, 524may have different cross-sectional dimensions (e.g., width, height orthickness) and shapes and may also be fabricated from differentmaterials.

In the illustrated embodiment, the crosstalk shields 522, 524 arecoupled to each other to form a single larger crosstalk shield. However,the crosstalk shields 522, 524 may be spaced apart from each other inother configurations. For example, the crosstalk shields 522, 524 may bespaced apart from each other along the longitudinal axis 468. In theillustrated embodiment, the crosstalk shields 522, 524 at leastpartially surround the input region 472 and a portion of the passivationlayer 454. The crosstalk shield 522 directly engages the shield layer450. In some embodiments, the crosstalk shields 522, 524 may onlypartially surround the light guide 462. In other embodiments, thecrosstalk shields 522, 524 may constitute crosstalk rings thatcircumferentially surround the entire light guide 462. Such embodimentsare described in greater detail below with respect to FIGS. 10 and 11.

As shown, the guide cavity 456 is defined by one or more interiorsurfaces 526 of the device base 425. In particular embodiments, theinterior surfaces 526 may be surface(s) of the dielectric material(e.g., SiO₂) from the substrate layers 432-437. The crosstalk shields522, 524 may directly abut the light guide such that a portion of themetallic elements is exposed to and directly engages the filter material460 of the light guide 462. In other embodiments, however, the crosstalkshields 522, 524 are not exposed to the light guide 462 and, instead,may be positioned immediately adjacent to the light guide 462 such thata portion of the dielectric material is located between the crosstalkshields 522, 524 and the light guide 462. For example, in theillustrated embodiment, dielectric material 528, 530 is located betweenthe light guide 462 and the crosstalk shields 522, 524, respectively.The dielectric material 528, 530 may each include a portion of theinterior surface 526. The dielectric material 528, 530 may separate thelight guide 462 from the respective crosstalk shields 522, 524 by aseparation distance SD. By way of example only, the separation distanceSD may be at least about 150 nm. In some embodiments, the separationdistance SD is at least about 100 nm. The separation distance SD may beless than 100 nm.

FIG. 10 is a schematic cross-section of a detection device 602 formed inaccordance with another embodiment. The detection device 602 may includesimilar features as the detection device 404 (FIG. 7) and may be used inbiosensors, such as the biosensor 400 (FIG. 7) or the biosensor 102(FIG. 1). The detection device 602 may also be manufactured usingintegrated circuit manufacturing technologies. The detection device 602is described and illustrated to demonstrate other features thatdetection devices and biosensors may have. In some embodiments, thedetection device 602 alone may constitute a biosensor. In otherembodiments, the detection device 602 may be coupled to a flow cell toform a biosensor. For example, the detection device 602 may be coupledto the flow cell 402 and form a flow channel between the detectiondevice 602 and the flow cell 402.

As shown, the detection device 602 includes a device base 604, a shieldlayer 640, and multiple sub-layers 652, 654 that collectively form apassivation layer 650 of the detection device 602. The device base 604includes a sensor array 606 of light sensors 608 and a guide array 610of light guides 612. The light sensors 608 may be similar or identicalto the light sensors 440, and the light guides 612 may be similar oridentical to the light guides 462. For example, the light guides 612 areconfigured to receive the excitation light 614 and the light emissions616. As shown, the light emissions 616 are illustrated as light beingemitted from a single point. It is understood that the light emissionsmay be generated from multiple points along the passivation layer 650.Each of the light guides 612 extends into the device base 604 along acentral longitudinal axis 618 from an input region 620 of the lightguide 612 toward a corresponding light sensor 608 of the sensor array606.

Similar to the light guides 462, the light guides 612 may include afilter material that is configured to filter the excitation light 614and permit the light emissions 616 to propagate therethrough toward thecorresponding light sensors 608. The device base 604 includes devicecircuitry (not shown) that is electrically coupled to the light sensors608 and configured to transmit data signals based on photons detected bythe light sensors. Although not shown in FIGS. 10 and 11, the circuitryof the device base 604 may be located between the light guides 612similar to the circuitry 446 (FIG. 8) located between the light guides462.

As shown, the device base 604 includes peripheral crosstalk shields631-634 that are located within the device base 604. More specifically,each of the light guides 612 is surrounded by multiple crosstalk shields631-634. The crosstalk shields 631-634 for each of the light guides 612may be spaced apart from each other along the respective longitudinalaxis 618 such that gaps 641-643 are formed therebetween. The sizes ofthe gaps 641-643 may be substantially equal to one another or maydiffer. For example, the gaps 643 are slightly larger than the gaps 642.

In the illustrated embodiment, the crosstalk shields 631-634 areconfigured to circumferentially surround the light guides 612. As usedherein, the phrase “circumferentially surround” is not intended torequire that the light guides 612 have circular cross-section and/or thecrosstalk shields 631-634 have circular shapes. Instead, a crosstalkshield may circumferentially surround the light guide 612 if thecrosstalk shield surrounds the corresponding longitudinal axis 618. Thecrosstalk shield may completely surround the longitudinal axis 618 oronly partially surround the longitudinal axis 618. For example, thecrosstalk shields 631-634 may continuously extend around thecorresponding light guide 612 or, in other cases, the crosstalk shields631-634 may include multiple sub-elements that are individuallydistributed around the light guide 612 to at least partially surroundthe corresponding light guide.

Similar to the shield layer 452, the shield layer 640 may form apertures642 therethrough. The apertures 642 are substantially aligned withcorresponding light guides 612 and light sensors 608 to permit lightsignals to propagate into the corresponding input regions 620. Thesub-layer 654 may be deposited over the shield layer 640 such that thematerial of the sub-layer 654 fills at least a portion of the apertures.In some embodiments, an additional sub-layer 652 is deposited over thesub-layer 654 to form the passivation layer 650. By way of example only,either of the sub-layers 652, 654 may include plasma vapor deposition(PVD) Ta₂O₅ or plasma-enhanced chemical vapor deposition (PECVD)Si_(x)N_(y). In another embodiment, an additional sub-layer may bestacked onto the sub-layers 652, 654. By way of one specific example,the sub-layer 654 may be PVD Ta₂O₅, the sub-layer 652 may be PECVDSi_(x)N_(y), and an additional layer that is stacked onto the sub-layer652 may be PVD Ta₂O₅.

FIG. 11 is a flowchart illustrating a method 700 of manufacturing abiosensor in accordance with one embodiment. The method 700 isillustrated in FIGS. 12A and 12B. The method 700, for example, mayemploy structures or aspects of various embodiments (e.g., systemsand/or methods) discussed herein. In various embodiments, certain stepsmay be omitted or added, certain steps may be combined, certain stepsmay be performed simultaneously, certain steps may be performedconcurrently, certain steps may be split into multiple steps, certainsteps may be performed in a different order, or certain steps or seriesof steps may be re-performed in an iterative fashion.

The method 700 may include providing (at 702) a device base 800 having asensor array of light sensors 802. As shown, the device base 800 has anouter or external surface 801. The device base 800 may be manufacturedusing integrated circuit manufacturing technologies, such as CMOSmanufacturing technologies. For example, the device base 800 may includeseveral substrate layers with different modified features (e.g.,metallic elements) embedded therein. In some embodiments, the devicebase 800 may include guide regions 804 and circuitry regions 806. Theguide regions 804 may correspond to portions of the device base 800 thatwill include, after the method 700, the light guides. Adjacent guideregions 804 may be separated by the circuitry regions 806 that includedevice circuitry (not shown), which may be similar to the devicecircuitry described herein. More specifically, the device circuitry maybe electrically coupled to the light sensors 802 and configured totransmit data signals based on photons detected by the light sensors802. In some embodiments, the guide regions 804 may include peripheralcrosstalk shields 808 that surround substrate material in the guideregions 804.

The method 700 may also include applying (at 704) a shield layer 810 tothe outer surface 801 of the device base 800 and forming (at 706)apertures 812 through the shield layer 810. As described above, theshield layer 810 may include a metal material that is configured toblock light signals. The apertures 812 may be formed by applying a mask(not shown) and removing material (e.g., through etching) of the shieldlayer 810 to form the apertures 812.

At 708, guide cavities 814 may be formed in the device base 800. Morespecifically, the substrate material within the guide regions 804 may beremoved so that the guide cavities 814 extend from proximate to theapertures 812 toward corresponding light sensors 802. As shown in FIG.12A, interior surfaces 815 of the substrate material may define theguide cavities 814. The guide cavities 814 may be sized and shaped suchthat the interior surfaces 815 are proximate to the crosstalk shields808. As described herein, the crosstalk shields 808 may be immediatelyadjacent to the interior surfaces 815 or may be exposed in the guidecavities 814.

The method 700 may also include depositing (at 710) filter material 820within the guide cavities 814. The filter material 820 may be, forexample, an organic filter material. In some embodiments, a portion ofthe filter material 820 may extend along the shield layer 810 after thedepositing operation. For example, the amount of the filter material 820applied to the device base 800 may exceed the available volume withinthe guide cavities 814. As such, the filter material 820 may overflowthe guide cavities 814 and extend along the shield layer 810.

In some embodiments, depositing (at 710) the filter material 820 mayinclude pressing (e.g., using a squeegee-like component) the filtermaterial 820 into the guide cavities 814. FIG. 12A appears to indicate auniform layer of the filter material 820 along the shield layer 810. Insome embodiments, the layer of filter material 820 may not be uniform.For instance, only portions of the shield layer 810 may have the filtermaterial 820 thereon. In alternative embodiments, the depositingoperation may include selectively filling each of the guide cavities 814such that the filter material 820 does not clear or overflow the guidecavities 814.

At 712, the filter material 820 may be cured. Optionally, the method 700may also include removing (at 714) the filter material 820 from theshield layer 810 and, in some cases, portions of the filter material 820from the guide cavities 814. The filter material 820 may be removed fromwithin the guide cavities 814 so that a material level 830 of the filtermaterial 820 is located within the aperture 812 or at a depth below theshield layer 810. In embodiments where the material level 830 is belowthe shield layer 810, the filter material 820 may not contact anymaterial of the shield layer 810. The filter material 820 within theguide cavities 814 may form light guides. Different processes may beimplemented for removing the filter material 820 from the shield layer810. For example, the removing operation may include at least one ofetching the filter material or chemically polishing the filter material.

As shown in FIG. 12B, the method 700 may also include applying (at 716)a passivation layer 832 to the shield layer 810 and to the filtermaterial 820 of the light guides such that the passivation layer 832extends directly along the shield layer 810 and across the apertures812. The passivation layer 832 may extend directly along the lightguides at corresponding material interfaces 834, such as the materialinterfaces 516 (FIG. 9). In the illustrated embodiment, the passivationlayer 832 has a planar detector surface 836. In other embodiments, thedetector surface 836 may form an array of reaction recesses, such as thereaction recesses 408 (FIG. 7). The reaction recesses may extend towardor be located within corresponding apertures 812.

In some embodiments, the passivation layer 832 includes multiplesub-layers 841-843. In particular embodiments, at least one of thesub-layers 841-843 includes tantalum. For example, the sub-layer 841 mayinclude tantalum pentoxide (Ta₂O₅), the sub-layer 842 may include alow-temperature film (e.g., silicon nitride (Si_(x)N_(y))), and thesub-layer 843, which may have the detector surface 836, may includetantalum pentoxide (Ta₂O₅). However, the sub-layers 841-843 are onlyprovided as examples and other passivation layers may include fewersub-layers, more sub-layers, or sub-layers with different materials. Insome cases, only a single sub-layer is used for the passivation layer.

Optionally, the method 700 may include providing (at 718) reaction sites850 and mounting a flow cell (not shown). Providing the reaction sites850 may occur prior to or after the flow cell is coupled to thedetection device. The reaction sites 850 may be located at designationaddresses such that the reaction sites 850 have a predetermined patternalong the detector surface 836. The reaction sites may correspond (e.g.,one site to one light sensor, one site to multiple light sensors, ormultiple sites to one light sensor) in a predetermined manner. In otherembodiments, the reaction sites may be randomly formed along thedetector surface 836. As described herein, the reaction sites 850 mayinclude biological or chemical substances immobilized to the detectorsurface 836. The biological or chemical substances may be configured toemit light signals in response to excitation light. In particularembodiments, the reaction sites 850 include clusters or colonies ofbiomolecules (e.g., oligonucleotides) that are immobilized on thedetector surface 836.

In an embodiment, a biosensor is provided that includes a flow cell anda detection device having the flow cell coupled thereto. The flow celland the detection device form a flow channel that is configured to havebiological or chemical substances therein that generate light emissionsin response to an excitation light. The detection device includes adevice base having a sensor array of light sensors and a guide array oflight guides. The light guides have input regions that are configured toreceive the excitation light and the light emissions from the flowchannel. The light guides extend into the device base from the inputregions toward corresponding light sensors and have a filter materialthat is configured to filter the excitation light and permit the lightemissions to propagate toward the corresponding light sensors. Thedevice base includes device circuitry electrically coupled to the lightsensors and configured to transmit data signals based on photonsdetected by the light sensors. The detection device also includes ashield layer that extends between the flow channel and the device base.The shield layer has apertures that are positioned relative to the inputregions of corresponding light guides such that the light emissionspropagate through the apertures into the corresponding input regions.The shield layer extends between adjacent apertures and is configured toblock the excitation light and the light emissions incident on theshield layer between the adjacent apertures.

In one aspect, the input regions of the light guides may be locatedwithin the corresponding apertures of the shield layer or may be locateda depth into the device base.

In another aspect, the detection device may include a passivation layerthat extends along the shield layer such that the shield layer isbetween the passivation layer and the device base. The passivation layermay extend across the apertures.

In particular cases, the filter material of the light guides may be anorganic filter material. The passivation layer may extend directly alongthe input regions of the light guides and isolate the organic filtermaterial from the shield layer. The material interfaces may be locatedwithin the corresponding apertures of the shield layer or located adepth into the device base. In certain embodiments, the passivationlayer extends into the apertures and forms an array of reactionrecesses. The reaction recesses may extend toward or be located withincorresponding apertures.

In certain embodiments, the biological or chemical substances areconfigured to be located within the reaction recesses. In certainembodiments, the reaction recesses have corresponding base surfaces. Thebase surfaces may be located within the aperture or located a depth intothe device base.

In another aspect, the device base includes peripheral crosstalkshields. Each of the crosstalk shields may surround one of thecorresponding light guides. The crosstalk shields may be configured toreduce optical crosstalk between adjacent light sensors.

In another aspect, the biosensor is lens-free such that the biosensordoes not include an optical element that focuses the light emissionstoward a focal point.

In an embodiment, a biosensor is provided that includes a flow cell anda detection device having the flow cell coupled thereto. The flow celland the detection device form a flow channel that is configured to havebiological or chemical substances therein that generate light emissionsin response to an excitation light. The detection device may include adevice base having a sensor array of light sensors and a guide array oflight guides. The light guides are configured to receive the excitationlight and the light emissions from the flow channel. Each of the lightguides extends into the device base along a central longitudinal axisfrom an input region of the light guide toward a corresponding lightsensor of the sensor array. The light guides include a filter materialthat is configured to filter the excitation light and permit the lightemissions to propagate therethrough toward the corresponding lightsensors. The device base includes device circuitry that is electricallycoupled to the light sensors and configured to transmit data signalsbased on photons detected by the light sensors. The device base includesperipheral crosstalk shields located therein that surround correspondinglight guides of the guide array. The crosstalk shields at leastpartially surround the corresponding light guides about the respectivelongitudinal axis to reduce optical crosstalk between adjacent lightsensors.

In one aspect, the crosstalk shields may surround the input regions ofthe corresponding light guides.

In another aspect, the crosstalk shields may include crosstalk ringsthat circumferentially surround the corresponding light guide.

In another aspect, the device base may include acomplementary-metal-oxide semiconductor (CMOS) and the device circuitry.The crosstalk shields may include metallic elements located withindielectric layers of the device base. The crosstalk shields may beelectrically separate from the device circuitry.

In another aspect, a shield layer may extend between the flow channeland the device base. The shield layer may have apertures that arepositioned relative to the input regions of corresponding light guidesof the guide array. The apertures may permit the light emissions topropagate therethrough into the input regions. The shield layer mayextend between adjacent apertures and is configured to block theexcitation light and the light emissions incident on the shield layerbetween the adjacent apertures. For instance, the input regions of thelight guides may be located within the corresponding apertures of theshield layer or are located a depth into the device base.

In another aspect, the detection device may also include a passivationlayer that extends along the shield layer such that the shield layer isbetween the passivation layer and the device base and across theapertures.

In another aspect, the crosstalk shield abuts or is immediately adjacentto the shield layer.

In another aspect, the crosstalk shields are first crosstalk shields,and the device base includes second crosstalk shields in which each ofthe light guides of the guide array is at least partially surrounded bycorresponding first and second crosstalk shields. For example, the firstand second crosstalk shields may be spaced apart from each other alongthe corresponding longitudinal axis. In another embodiment, the firstand second crosstalk shields have different dimensions.

In an embodiment, a method of manufacturing a biosensor is provided. Themethod includes providing a device base having a sensor array of lightsensors and device circuitry that is electrically coupled to the lightsensors and configured to transmit data signals based on photonsdetected by the light sensors. The device base has an outer surface. Themethod also includes applying a shield layer to the outer surface of thedevice base and forming apertures through the shield layer. The methodalso includes forming guide cavities that extend from correspondingapertures toward a corresponding light sensor of the sensor array anddepositing filter material within the guide cavities. A portion of thefilter material extends along the shield layer. The method also includescuring the filter material and removing the filter material from theshield layer. The filter material within the guide cavities forms lightguides. The method also includes applying a passivation layer to theshield layer such that the passivation layer extends directly along theshield layer and across the apertures.

In one aspect, removing the filter material from the shield layerincludes removing a portion of the filter material within the guidecavities such that a material level of the filter material is locatedwithin the aperture or at a depth below the shield layer.

In another aspect, the passivation layer extends directly along thelight guides at corresponding material interfaces. The materialinterfaces are located within the corresponding apertures or located adepth into the device base.

In another aspect, the filter material is an organic filter material.The passivation layer extends directly along the light guides andisolates the organic filter material from the shield layer.

In another aspect, the passivation layer forms an array of reactionrecesses. The reaction recesses extend toward or are located withincorresponding apertures. For instance, the reaction recesses may havecorresponding base surfaces. The base surfaces may be located within theaperture or located a depth into the device base.

In another aspect, the method includes coupling a flow cell to thedevice base to form a flow channel between the passivation layer and theflow cell.

In another aspect, removing the filter material from the shield layerincludes at least one of etching the filter material or chemicallypolishing the filter material.

In another aspect, the passivation layer includes tantalum pentoxide(Ta₂O₅). For example, the passivation layer may include multiplesub-layers in which at least one of the sub-layers includes tantalumpentoxide (Ta₂O₅). In a more specific embodiment, the sub-layers mayinclude two tantalum pentoxide layers with a low-temperature filmtherebetween.

In another aspect, the device base has guide regions that includesubstrate material prior to forming the guide cavities in which adjacentguide regions are separated by circuitry regions that include the devicecircuitry. Forming the guide cavities may include removing the substratematerial of the guide regions.

In another aspect, the device base may include peripheral crosstalkshields that surround the guide regions prior to forming the guidecavities. The crosstalk shields may at least partially surround thecorresponding light guides after the light guides are formed. Thecrosstalk shields may be configured to reduce optical crosstalk betweenadjacent light sensors.

In an embodiment, a biosensor is provided that includes a device basehaving a sensor array of light sensors and a guide array of lightguides. The device base has an outer surface. The light guides haveinput regions that are configured to receive excitation light and lightemissions generated by biological or chemical substances proximate tothe outer surface. The light guides extend into the device base from theinput regions toward corresponding light sensors and have a filtermaterial that is configured to filter the excitation light and permitthe light emissions to propagate toward the corresponding light sensors.The device base includes device circuitry electrically coupled to thelight sensors and configured to transmit data signals based on photonsdetected by the light sensors. The biosensor also includes a shieldlayer that extends along the outer surface of the device base. Theshield layer has apertures that are positioned relative to the inputregions of corresponding light guides such that the light emissionspropagate through the apertures into the corresponding input regions.The shield layer extends between adjacent apertures and is configured toblock the excitation light and the light emissions incident on theshield layer between the adjacent apertures.

In an embodiment, a biosensor is provided that includes a device basehaving a sensor array of light sensors and a guide array of lightguides. The device base has an outer surface. The light guides areconfigured to receive excitation light and light emissions generated bybiological or chemical substances proximate to the outer surface. Eachof the light guides extends into the device base along a centrallongitudinal axis from an input region of the light guide toward acorresponding light sensor of the sensor array. The light guide includesa filter material that is configured to filter the excitation light andpermit the light emissions to propagate therethrough towardcorresponding light sensors. The device base includes device circuitrythat is electrically coupled to the light sensors and are configured totransmit data signals based on photons detected by the light sensors.The device base includes peripheral crosstalk shields located thereinthat surround corresponding light guides of the guide array. Thecrosstalk shields at least partially surrounding the corresponding lightguides about the respective longitudinal axis to at least one of blockor reflect errant light rays to reduce optical crosstalk betweenadjacent light sensors.

It is to be understood that the subject matter described herein is notlimited in its application to the details of construction and thearrangement of components set forth in the description herein orillustrated in the drawings hereof. The subject matter described hereinis capable of other embodiments and of being practiced or of beingcarried out in various ways. Also, it is to be understood that thephraseology and terminology used herein is for the purpose ofdescription and should not be regarded as limiting. The use of“including,” “comprising,” or “having” and variations thereof herein ismeant to encompass the items listed thereafter and equivalents thereofas well as additional items.

Unless specified or limited otherwise, the terms “mounted,” “connected,”“supported,” and “coupled” and variations thereof are used broadly andencompass both direct and indirect mountings, connections, supports, andcouplings. Further, “connected” and “coupled” are not restricted tophysical or mechanical connections or couplings. Also, it is to beunderstood that phraseology and terminology used herein with referenceto device or element orientation (such as, for example, terms like“above,” “below,” “front,” “rear,” “distal,” “proximal,” and the like)are only used to simplify description of one or more embodimentsdescribed herein, and do not alone indicate or imply that the device orelement referred to must have a particular orientation. In addition,terms such as “outer” and “inner” are used herein for purposes ofdescription and are not intended to indicate or imply relativeimportance or significance.

It is to be understood that the above description is intended to beillustrative, and not restrictive. For example, the above-describedembodiments (and/or aspects thereof) may be used in combination witheach other. In addition, many modifications may be made to adapt aparticular situation or material to the teachings of the presentlydescribed subject matter without departing from its scope. While thedimensions, types of materials and coatings described herein areintended to define the parameters of the disclosed subject matter, theyare by no means limiting and are exemplary embodiments. Many otherembodiments will be apparent to those of skill in the art upon reviewingthe above description. The scope of the inventive subject matter should,therefore, be determined with reference to the appended claims, alongwith the full scope of equivalents to which such claims are entitled. Inthe appended claims, the terms “including” and “in which” are used asthe plain-English equivalents of the respective terms “comprising” and“wherein.” Moreover, in the following claims, the terms “first,”“second,” and “third,” etc. are used merely as labels, and are notintended to impose numerical requirements on their objects. Further, thelimitations of the following claims are not written inmeans—plus-function format and are not intended to be interpreted basedon 35 U.S.C. § 112, sixth paragraph, unless and until such claimlimitations expressly use the phrase “means for” followed by a statementof function void of further structure.

The following claims recite aspects of certain embodiments of theinventive subject matter and are considered to be part of the abovedisclosure. These aspects may be combined with one another.

What is claimed is:
 1. A device comprising: a flow cell; and a detectiondevice having the flow cell coupled thereto, the flow cell and thedetection device forming a flow channel that is configured to havebiological or chemical substances therein that generate light emissionsin response to an excitation light, the detection device including: adevice base having a sensor array of light sensors and a guide array oflight guides, the device base having an outer surface, the light guidesconfigured to receive the excitation light and the light emissions fromthe flow channel, each of the light guides extending into the devicebase along a central longitudinal axis from an input region of the lightguide toward a corresponding light sensor of the sensor array, the lightguides including a filter material that is configured to filter theexcitation light and permit the light emissions to propagatetherethrough toward the corresponding light sensors, the device baseincluding device circuitry that is electrically coupled to the lightsensors and configured to transmit data signals based on photonsdetected by the light sensors; a passivation layer that extends over theouter surface of the device base and forms an array of reaction recessesabove the light guides; wherein the device base includes peripheralcrosstalk shields located therein that at least partially surroundcorresponding light guides of the guide array, the crosstalk shields atleast partially surrounding the corresponding light guides about therespective longitudinal axis to reduce optical crosstalk betweenadjacent light sensors.
 2. The device of claim 1, wherein the crosstalkshields surround the input regions of the corresponding light guides. 3.The device of claim 1, wherein the crosstalk shields include crosstalkrings that circumferentially surround the corresponding light guide. 4.The device of claim 1, wherein the device base includes acomplementary-metal-oxide semiconductor (CMOS) and the crosstalk shieldsinclude metallic elements located within dielectric layers of the devicebase, the crosstalk shields being electrically separate from the devicecircuitry.
 5. The device of claim 1, wherein the reaction recesses havecorresponding base surfaces, the base surfaces being located above thelight guide at a depth into the device base.
 6. The device of claim 1,further comprising a shield layer extending between the passivationlayer and the device base, the shield layer having apertures that arepositioned relative to the input regions of corresponding light guidesof the guide array, the apertures permitting the light emissions topropagate therethrough into the input regions, the shield layerextending between adjacent apertures and configured to block theexcitation light and the light emissions incident on the shield layerbetween the adjacent apertures.
 7. The device of claim 6, wherein theinput regions of the light guides are located within the correspondingapertures of the shield layer or are located a depth into the devicebase.
 8. The device of claim 6, wherein the passivation layer extendsalong the shield layer such that the shield layer is between thepassivation layer and the device base, the passivation layer extendingacross the apertures.
 9. The device of claim 8, wherein the passivationlayer includes tantalum pentoxide (Ta₂O₅).
 10. The device of claim 8,wherein the passivation layer includes multiple sub-layers in which atleast one of the sub-layers includes tantalum pentoxide (Ta₂O₅).
 11. Thedevice of claim 10, wherein the sub-layers includes two tantalumpentoxide layers with a low-temperature film therebetween.
 12. Thedevice of claim 10, wherein the sub-layers include two plasma vapordeposition (PVD) tantalum pentoxide layers with a plasma enhancedchemical vapor deposition (PECVD) silicon nitride film therebetween. 13.The device of claim 6, wherein the crosstalk shields abut or areimmediately adjacent to the shield layer.
 14. The device of claim 6,wherein the shield layer is in contact with the passivation layer, butis not in contact with the filter material of the light guides.
 15. Thedevice of claim 1, wherein the crosstalk shields are first crosstalkshields, the device base including second crosstalk shields, whereineach of the light guides of the guide array is at least partiallysurrounded by corresponding first and second crosstalk shields.
 16. Thedevice of claim 15, wherein the first and second crosstalk shields arespaced apart from each other along the corresponding longitudinal axis.17. The device of claim 15, wherein the first and second crosstalkshields have different dimensions.
 18. A device comprising: a devicebase having a sensor array of light sensors and a guide array of lightguides, the device base having an outer surface, the light guides havinginput regions that are configured to receive excitation light and lightemissions generated by biological or chemical substances proximate tothe outer surface, the light guides extending into the device base fromthe input regions toward corresponding light sensors and having a filtermaterial that is configured to filter the excitation light and permitthe light emissions to propagate toward the corresponding light sensors,the device base including device circuitry electrically coupled to thelight sensors and configured to transmit data signals based on photonsdetected by the light sensors; wherein the device base includesperipheral crosstalk shields located therein that surround correspondinglight guides of the guide array, the crosstalk shields at leastpartially surrounding the corresponding light guides about therespective longitudinal axis to at least one of block or reflect errantlight rays to reduce optical crosstalk between adjacent light sensors.19. The device of claim 18, further comprising a passivation layer thatextends over the outer surface of the device base and forms an array ofreaction recesses above the light guides.
 20. The device of claim 19,wherein the reaction recesses have corresponding base surfaces, the basesurfaces being located above the light guide at a depth into the devicebase.
 21. A method of manufacturing a device, the method comprising:forming guide cavities in a device base, the device base having a sensorarray of light sensors and device circuitry that is electrically coupledto the light sensors and to transmit data signals based on photonsdetected by the light sensors, the device base having an outer surfaceand peripheral crosstalk shields extending from the outer surface towardthe light sensors; wherein the guide cavities extend from correspondingapertures toward a corresponding light sensor of the sensor array, suchthat the guide cavities are separated by the peripheral crosstalkshields; depositing filter material within the guide cavities, thefilter material within the guide cavities forming light guides; curingthe filter material; and applying a passivation layer over the devicebase that extends over the light guides.
 22. The method of claim 21,further comprising applying a shield layer to the outer surface of thedevice base prior to applying the passivation layer, and formingapertures through the shield layer between the peripheral crosstalkshields, wherein the passivation layer extends directly along the shieldlayer and across the apertures.